Simplified Direct PCR Method for Reference Buccal Samples Using a Non-FTA Card by Omitting the Pretreatment Step

When using non-FTA cards in commercial multiplex STR kits for direct PCR, pretreatment steps with specific buffers are recommended. Here, we designed a rapid direct PCR method utilizing a non-FTA card, Oral Cell Sampling Kit, by omitting the pretreatment step involving Prep-n-Go™ Buffer, and it showed compatibility with the GlobalFiler™ Express PCR Amplification Kit, GlobalFiler™ PCR Amplification Kit, and PowerPlex^® Fusion system. To optimize the PCR conditions, we tested the method with different final PCR volumes and cycles. Finally, we conducted a performance test using 50 Korean buccal samples and confirmed the high performance of the method, detecting more than 90% of the samples with full profiles when using GlobalFiler™ PCR Amplification Kit and PowerPlex^® Fusion system at 29 cycles in a 10 μL final PCR volume. Thus, we report a simple direct PCR set-up to analyze reference samples collected using a non-FTA card manufactured in Korea.     

Whole Plastome Sequences of Two Drug-Type Cannabis: Insights Into the Use of Plastid in Forensic Analyses

DNA is one of the fastest growing tools in forensic sciences, increasing reliability in forensic reports and judgments. The use of DNA has increased in different areas of the forensic sciences, such as investigation of plant species, where plastid DNA has been used to elucidate and generate evidence in cases of traceability of genetically modified and controlled plants. Even with several advances and the practice of using DNA in forensic investigations, there are just few studies related to the identification of genetic tools for the characterization of drug and nondrug-types of Cannabis. Herein, the whole plastomes of two drug-type Cannabis are presented and have their structures compared with other Cannabis plastomes deposited in the GenBank, focusing in the forensic use of plastome sequences. The plastomes of Cannabis sativa “Brazuka” and of the hybrid Cannabis AK Royal Automatic presented general structure that does not differs from the reported for other C. sativa cultivars. A phylogenomic analyses grouped C. sativa “Brazuka” with the nondrug C. sativa cultivars, while the hybrid Cannabis AK Royal Automatic placed isolated, basal to this group. This suggests that the analysis of plastomes is useful toward genetic identification of hybrids in relation to C. sativa.     

The Effect of Temperature and Exposure Time on Stability of Cholesterol and Squalene in Latent Fingermarks Deposited on PVDF Membrane

Cholesterol and squalene are fatty materials of latent fingermarks that can be utilized for dating methodologies and visualization techniques. Previous studies have suggested these compounds undergo degradation in fingermarks as a function of time (days) and light at ambient temperature. However, studies assessing how their composition changes at low and high temperatures over short periods of time (hours) have not been published previously. Here, we performed quantitative analysis of cholesterol and squalene in natural fingermark residue using PVDF membrane, after exposure to a range of temperatures (−20 to 100°C) for 4 and 8 h. We found that levels of both fatty materials remained constant at −20 to 60°C, but both showed significant reduction at 100°C, over short exposure times. These results indicate that cholesterol and squalene are detectable at −20 to 60°C, whereas at 100°C or higher, both are lost due to rapid thermal degradation.     

Evaluating Bioinformatic Pipeline Performance for Forensic Microbiome Analysis*,†,‡

Microbial communities have potential evidential utility for forensic applications. However, bioinformatic analysis of high-throughput sequencing data varies widely among laboratories. These differences can potentially affect microbial community composition and downstream analyses. To illustrate the importance of standardizing methodology, we compared analyses of postmortem microbiome samples using several bioinformatic pipelines, varying minimum library size or minimum number of sequences per sample, and sample size. Using the same input sequence data, we found that three open-source bioinformatic pipelines, MG-RAST, mothur, and QIIME2, had significant differences in relative abundance, alpha-diversity, and beta-diversity, despite the same input data. Increasing minimum library size and sample size increased the number of low-abundant and infrequent taxa detected. Our results show that bioinformatic pipeline and parameter choice affect results in important ways. Given the growing potential application of forensic microbiology to the criminal justice system, continued research on standardizing computational methodology will be important for downstream applications.     

Unusual and Fatal Case of an Undiagnosed Intravascular Large B-cell Lymphoma: The Oncologist’s Great Imitator†

Intravascular lymphoma (IVL) is a rare subtype of extranodal lymphomas that is characterized by the selective growth of neoplastic cells within the lumen of small vessels. Authors document the case of an unexpected death caused by an undiagnosed intravascular large B-cell lymphoma with multi-organ involvement, which had initially manifested as an infection and then as an unclarified central nervous system pathology. Histological examination showed a diffuse intravascular large B-cell brain lymphoma with prominent cerebral involvement. The relevance of the case report reveals the importance of an autopsy of an extremely rare and threatening pathology that in most cases is diagnosed only postmortem. As a result, the role of the forensic pathologist becomes particularly important. When specifically performing an in-depth autopsy evaluation with a specific histologic analysis, it is possible to identify the intravascular lymphoma and declare a more accurate cause of death.     

Ethyl Glucuronide Concentration in Hair of Detainees: A Preliminary Study

Through the measurement of ethyl glucuronide in hair (hETG), it is possible to assess chronic alcohol abuse over time. In this paper, we present a study on hETG in Italian prison inmates. Analyses were performed by LC-MS according to a previously published method. Results were evaluated using the cut-offs established by the Society of Hair Testing. Positives samples (ETG > 30 pg/mg) accounted for 6% of all subjects, with concentrations ranging from 42 pg/mg up to 270 pg/mg, abstinent subjects (ETG < 7 pg/mg) accounted for 88%, and moderate alcohol consumption (7 < ETG < 30 pg/mg) for 6% of the subjects. No females displayed ETG values above 30 pg/mg. Among positive samples, only two subjects did not declare heavy alcohol consumption and were found strongly positive at 210 and 270 pg/mg. To the best of our knowledge, this represents the first study on ETG hair concentration on prison inmates.     

A Comparison of Coronary Artery Stenosis Estimates Made by Forensic Pathologists and Medical Students

Previous studies suggest cardiovascular pathologists are less accurate than noncardiovascular pathologists (e.g., clinical pathologists) in estimating the degree of coronary artery stenosis. To further investigate the effect of training on accurate estimation of coronary artery stenosis, we designed a study to compare the accuracy of estimates made by forensic pathologists versus medical students. Six forensic pathologists and twelve medical students each independently examined 24 images of coronary artery cross sections and gave an estimate of the degree of stenosis. When comparing all 24 images, the forensic pathologists had a median difference between the estimated percentage of stenosis and actual percentage of stenosis of −12.380 and the medical students had a median difference of −16.50 (p-value of 0.08542). In estimating the percentage of stenosis, training in forensic pathology does not guarantee significantly improved accuracy compared with medical students. Our study showed no consistent statistically significant difference between estimates given by forensic pathologists and by medical students.     

A Study of the Background Levels of Male DNA on Underpants Worn by Females

This study is the first to examine the background level of male DNA on underpants worn by females in the absence of sexual contact. Here, we examined 103 samples from the inside front of underpants from 85 female volunteers. Samples were examined for the presence of male DNA using NGM SElect and PowerPlex Y23 kits. Only five samples gave a “complete” Y-STR profile, even though 83.5% of our volunteers cohabited with a male. In all cases where a partner reference sample was available, the Y-STR profile matched the cohabiting partner. We have demonstrated that a Y-STR profile is not expected on the inside front of underpants worn by females after social contact alone. The results of this study are informative for evaluating the significance of a Y-STR profile on underpants in cases of alleged sexual assault.     

Sample Mining and Data Mining: Combined Real-Time and Retrospective Approaches for the Identification of Emerging Novel Psychoactive Substances

Novel psychoactive substances (NPS) are synthetic drugs that pose serious public health and safety concerns. A multitude of NPS have been identified in the United States, often implicated in forensic investigations. The most common and effective manner for identifying NPS is by use of mass spectrometry and the true utility lies within nontargeted acquisition techniques. During this study, a liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) assay was developed, validated, and implemented for forensic toxicology testing. A SCIEX TripleTOF™ 5600 + with SWATH^® acquisition was used. Resulting data were compared against an extensive library database containing more than 800 compounds. The LC-QTOF-MS assay was applied to the reanalysis of biological sample extracts to discover emergent NPS. More than 3,000 sample extracts were analyzed, and more than 20 emerging NPS were detected for the first time. Among these were isopropyl-U-47700, 3,4-methylenedioxy-U-47700, fluorofuranylfentanyl, N-methyl norfentanyl, 2F-deschloroketamine, 3,4-methylenedioxy-alpha-PHP, eutylone, and N-ethyl hexedrone.