Assessing the feasibility of free DNA for disaster victim identification and forensic applications.

In tropical disaster victim identification (DVI) scenarios, challenging environmental conditions lead to accelerated DNA degradation in remains. To further enhance the utilization of leached DNA from tissue in the preservative solution (termed "free DNA") as an alternative source, we incorporated new results by assessing its integrity in postmortem and decomposing cadavers preserved in DNA/RNA Shield™ and modified TENT, with silica-based purification (QIAquick®) for faster processing. The psoas muscle tissues of one decomposed and ten cadavers were preserved in each solution at 25 °C and 35 °C for 3 months. Free DNA efficiency was compared with individual reference samples for reliable results in quantity, quality, and STR profiles. The findings revealed that DNA/RNA Shield™ effectively preserves free DNA integrity for extended storage, while modified TENT is more suitable for short-term storage due to higher degradation levels. Moreover, the use of free DNA samples with massive parallel sequencing displays potential for forensic DNA analysis. Successful amplification of the mtDNA control region enables variant calling and heteroplasmy analysis while also serving as quality control using ACTB and enabling differentiation within the 16S rRNA region for microbiome analysis. The simplicity of handling free DNA for PCR-based forensic analysis adds to its potential for various applications, including DVI and field-based analysis of biological evidence.

Unraveling the mitochondrial phylogenetic landscape of Thailand reveals complex admixture and demographic dynamics.

The evolutionary dynamics of mitochondrial DNA within the Thai population were comprehensively explored with a specific focus on the influence of South Asian admixture. A total of 166 samples were collected through randomized sampling, ensuring a diverse representation. Our findings unveil substantial genetic and haplogroup diversity within the Thai population. We have identified 164 haplotypes categorized into 97 haplogroups, with a notable inclusion of 20 novel haplogroups. The distribution of haplogroups exhibited variations across different populations and countries. The central Thai population displayed a high diversity of haplogroups from both the M and N clades. Maternal lineage affinities were discerned between several Mainland Southeast Asia (MSEA) and South Asian populations, implying ancestral genetic connections and a substantial influence of South Asian women in establishing these relationships. f4-statistics indicates the presence of a Tibeto-Burman genetic component within the Mon population from Thailand. New findings demonstrate two phases of population expansion occurring 22,000-26,000 and 2500-3800 years ago, coinciding with the Last Glacial Maximum, and Neolithic demographic transition, respectively. This research significantly enhances our understanding of the maternal genetic history of Thailand and MSEA, emphasizing the influence of South Asian admixture. Moreover, it underscores the critical role of prior information, such as mutation rates, within the Bayesian framework for accurate estimation of coalescence times and inferring demographic history.

Levodopa-induced dyskinesia in early-onset Parkinson's disease (EOPD) associates with glucocerebrosidase mutation: A next-generation sequencing study in EOPD patients in Thailand.

BACKGROUND: With the benefit of using next-generation sequencing (NGS), our aim was to examine the prevalence of known monogenic causes in early-onset Parkinson's disease (EOPD) patients in Thailand. The association between clinical features, such as levodopa-induced dyskinesia (LID), and genotypes were also explored. METHOD: NGS studies were carried out for EOPD patients in the Tertiary-referral center for Parkinson's disease and movement disorders. EOPD patients who had LID symptoms were enrolled in this study (n = 47). We defined EOPD as a patient with onset of PD at or below 50 years of age. LID was defined as hyperkinetic movements including chorea, ballism, dystonia, myoclonus, or any combination of these movements resulting from levodopa therapy, which could be peak-dose, off-period, or diphasic dyskinesias. RESULTS: Pathogenic variants were identified in 17% (8/47) of the Thai EOPD patients, of which 10.6% (5/47) were heterozygous GBA variants (c.1448T>C in 3 patients and c.115+1G>A in 2 patients), 4.3% (2/47) homozygous PINK1 variants (c.1474C>T) and 2.1% (1/47) a PRKN mutation (homozygous deletion of exon 7). The LID onset was earlier in patients with GBA mutations compared to those without (34.8±23.4 vs 106.2±59.5 months after starting levodopa, respectively, p = 0.001). LID onset within the first 30 months of the disease was also found to be independently associated with the GBA mutation (odds ratio [95% confidence interval] = 25.00 [2.12-295.06], p = 0.011). CONCLUSION: Our study highlights the high prevalence of GBA pathogenic variants in Thai patients with EOPD and the independent association of these variants with the earlier onset of LID. This emphasizes the importance of genetic testing in this population.

STRategy: A support system for collecting and analyzing next-generation sequencing data of short tandem repeats for forensic science.

Short tandem repeats (STRs) are short repeated sequences commonly found in the human genome and valuable in forensic science, used for human identity and relatedness markers. Next-generation sequencing (NGS) technologies, e.g., ForenSeq Signature Prep, can sequence STRs, inferring length-based alleles and single nucleotide polymorphisms (SNPs) and providing valuable insights into population and sub-population structures. Despite the potential benefits of NGS for STRs, no open-source software platform integrates the collection, management, and analysis of STR data from NGS into one place. Users must use multiple programs to process their STR data and then collect the results into a separate database or a file system folder. Moreover, analyzing repeat structures (STR repeat motifs) may require learning multiple software tools, making the process inefficient and cumbersome. To address this gap, we introduce the STRategy, a standalone web-based application supporting essential STR data management and analysis capabilities. The STRategy allows users to collect their data into its database, automatically calculates forensic parameters, and visualizes the analyzed data in various forms. Users can search the database using different options, such as by profile, loci, and genotypes, with and without a specific test kit. Moreover, users can also find the nucleotide variants of a locus among the samples. We designed the STRategy for internal use in a laboratory or an organization. Hence, our system includes role-based access control that allows users to search for or access specific data based on their responsibilities. The administrator role can customize the system, for example, configure maps according to the samples' geographic data, and manage reference STR repeat motifs. A laboratory or an organization can download and install a copy of STRategy on their local system using Docker, as described in https://github.com/cucpbioinfo/STRategy. In summary, the STRategy is an end-to-end system that provides users with a database to collect the analyzed STR data from NGS, the dynamic analyses of forensic parameters, and the variants of STR patterns according to the newly added samples, which are then explorable via various search options and visualizations. The system is helpful for both forensic investigations and forensic genetics.

The concordance of length- and sequence-based STRs used in forensic markers with guidance for practice.

Next-generation sequencing (NGS) technology has shed light on every aspect of genetic discoveries, including forensic genetics. The Miseq® FGx Forensic Genetic System (Verogen) is one of the pioneering forensic NGS that provided a complete system from library preparation to data analysis. The system has been validated by several studies and led to a more practical aspect. Short tandem repeat (STR) is a well-established marker that was designed specifically for human individualization. Since NGS provides different data from fragment analysis, a new STR nomenclature is established to make NGS backward compatible with the previous data. In this study, Thai population were used to evaluate the Miseq® FGx Forensic genetic system (Verogen) in practical aspect, including concordance study and forensic population parameters. In summary, we purposed a practical guideline for sequence-based STRs.

A minimal number CpGs of ELOVL2 gene for a chronological age estimation using pyrosequencing.

Chronological age estimation is an important piece of human identification used in forensic practice. Epigenetic modifications, especially DNA methylation, have been proposed to predict age. The methylation of the ELOVL2 gene is one of the age-related markers that could be tested in fresh or postmortem blood sample. We study the use of DNA methylation markers on the ELOVL2 gene and develop a prediction model to estimate the age from a postmortem blood sample using pyrosequencing. From 100 anonymous blood samples, a correlation study of DNA methylation and age was investigated. The regression analysis revealed 2 CpG sites for model prediction with an adjusted R2 value of 0.7 (p < 0.01). The model explained 74% of the variation in postmortem blood samples (n = 36) with a prediction error (RMSE) of 10.2 years and a mean absolute deviation (MAD) of 7.1 years, whereas the model (excluding a younger age group) had improved with a RMSE of 5.6 years and a MAD of 4.2 years. The performance parameters were analyzed in several simulated models and indicated that these markers are advantageous for age estimation in forensic scenarios. Finally, a robustness and reproducibility of the pyrosequencing technique would enable this approach to be the part of an age prediction in forensic investigation.

Determination of morphologic and immunohistochemical stain (p57 kip2) discrepancy of complete and partial hydatidiform mole by using microsatellite genotyping.

OBJECTIVE: to evaluate the role of microsatellite genotyping in discordant results between morphologic examination and p57Kip2 staining in hydatidiform mole. MATERIALS AND METHODS: 127 cases of hydatidiform mole who had morphologic examination and p57Kip2immunohistochemical staining were evaluated. Six discrepant cases between morphologic examination and p57Kip2 staining were recruited. DNA was extracted from chorionic villi and paired maternal decidual tissue in Formalin fixed paraffin embedded tissue sections. The STR DNA genotyping was performed by Applied Biosystems 3500 Genetic Analyzer. Genetic data analysis was performed by Gene mapper ID-X software. Three concordant cases were used as control. Results were compared to histopathology, p57Kip2 stain and development of post-molar GTN. RESULTS: All controlled cases were confirmed PHM. Two cases of histologic CHM with positive p57Kip2and 2 cases of PHM with negative p57Kip2 were reported as PHM from microsatellite. Other 2 cases of histologic diagnosis PHM with negative p57Kip2 reported as CHM from microsatellite test and both of them developed post-molar GTN. CONCLUSION: Microsatellite genotyping is a high accuracy method for differential diagnosis from complete and partial hydatidiform moles. However, cost of microsatellite genotyping is still too high to use routinely. Therefore, selected use in discrepancy cases may be suitable.

A SNP panel for early detection of artificial chimerism in HSCT patients using TaqMan technology.

The monitoring of chimerism status in a hematopoietic stem cell transplantation patient is a crucial process and is performed periodically in a short time interval. A short tandem repeat marker is widely used for chimerism analysis due to its high discrimination power. However, the sensitivity of this approach was limited to 5% of a minor contributor and the interpretation is usually interrupted with PCR stochastic phenomena. Here, we developed an SNP panel for chimerism analysis using TaqMan technology. A set of SNPs was selected from Thai ancestry informative markers and open-access databases with proper criteria. We examined the 30 recipient-donor pairs that underwent HSCT and showed that the panel can provide an informative marker from 90% of all pairs. An early detection of artificial chimerism in post-HSCT samples was observed when compared with STR analysis. In addition, the detail of cases was discussed.

Genetic variation study of 12 X chromosomal STR in central Thailand population.

Genetic data from 12 short tandem repeats (STR) on the X chromosome are currently used in forensics studies to resolve issues related to complex kinship or when data is missing or ambiguous. In this study, we genotyped these 12 X chromosome STR in DNA collected from individuals from central Thailand (n = 391, 282 men and 109 women) and used this information to calculate allele and haplotype frequencies as well as forensic parameters for kinship calculations. Polymorphism information contents of the loci were range from 0.5283-0.9247, and powers of discrimination in females and males were 0.7666-0.9905 and 0.6085-0.9291, respectively. A diallelic pattern was observed at the locus DXS7132. Moreover, a comparison of genetic distance revealed a close relationship within Asian countries. Our results indicate that the X chromosomal short tandem repeat (X-STR) multiplex system provides highly informative genetic data and could be advantageous in forensic studies.

Ancestry informative markers for distinguishing between Thai populations based on genome-wide association datasets.

The main purpose of this work was to identify a set of AIMs that stratify the genetic structure and diversity of the Thai population from a high-throughput autosomal genome-wide association study. In this study, more than one million SNPs from the international HapMap database and the Thai depression genome-wide association study have been examined to identify ancestry informative markers (AIMs) that distinguish between Thai populations. An efficient strategy is proposed to identify and characterize such SNPs and to test high-resolution SNP data from international HapMap populations. The best AIMs are identified to stratify the population and to infer genetic ancestry structure. A total of 124 AIMs were clearly clustered geographically across the continent, whereas only 89 AIMs stratified the Thai population from East Asian populations. Finally, a set of 273 AIMs was able to distinguish northern from southern Thai subpopulations. These markers will be of particular value in identifying the ethnic origins in regions where matching by self-reports is unavailable or unreliable, which usually occurs in real forensic cases.

Pseudomonas aeruginosa septicemia causes death following liposuction with allogenic fat transfer and gluteal augmentation.

Cosmetic surgery to improve aesthetic and body conditions is becoming increasingly popular worldwide. In 2013, the American Society of Plastic Surgeons (ASPS) reported that one of the top five cosmetic procedures in the US is liposuction with over 200,000 procedures per year. This type of surgery is regarded as a minimal risk operation. Since surgical complications are not often reported, liposuction is usually performed in outpatient clinics. Fatality after cosmetic liposuction surgery is also relatively rare. This case report presents a death following cosmetic liposuction with allogenic fat transfer and gluteal augmentation. The medico-legal autopsy, pathology, and postmortem microbiology examinations reveal that septicemia by Pseudomonas aeruginosa was the definite cause of death. Surgical risk assessment and pathogenesis of the organism was reviewed.