Volatile Organic Compound Profiling from Postmortem Microbes using Gas Chromatography–Mass Spectrometry

Volatile organic compounds (VOCs) are by-products of cadaveric decomposition and are responsible for the odor associated with decomposing remains. The direct link between VOC production and individual postmortem microbes has not been well characterized experimentally. The purpose of this study was to profile VOCs released from three postmortem bacterial isolates (Bacillus subtilis, Ignatzschineria indica, I. ureiclastica) using solid-phase microextraction arrow (SPME Arrow) and gas chromatography–mass spectrometry (GC-MS). Species were inoculated in headspace vials on Standard Nutrient Agar and monitored over 5 days at 24°C. Each species exhibited a different VOC profile that included common decomposition VOCs. VOCs exhibited upward or downward temporal trends over time. Ignatzschineria indica produced a large amount of dimethyldisulfide. Other compounds of interest included alcohols, aldehydes, aromatics, and ketones. This provides foundational data to link decomposition odor with specific postmortem microbes to improve understanding of underlying mechanisms for decomposition VOC production.     

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.     

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.     

Detection of Mitragynine in Mitragyna Speciosa (Kratom) Using Surface-Enhanced Raman Spectroscopy with Handheld Devices

A simple, quick, selective, sensitive, and effective field-friendly method capable of being used by nonexperts has been developed for detecting mitragynine in Mitragyna speciosa (kratom) using surface-enhanced Raman spectroscopy (SERS). Over 100 samples and blanks (known to be either positive or negative for the presence of mitragynine) were examined in duplicate using five identical handheld Raman spectrometers, which provided a data set of over 1,000 examinations. Based on the results of these analyses, the method yielded a true-positive rate of 99.3%, a true-negative rate of 97.9%, a false-positive rate of 2.1%, and a false-negative rate of 0.7%. The average minimum detectable concentration (C_m) of mitragynine that reproducibly yielded a match for one of the library spectra on all five instruments was determined to be 342 ng/mL (ppb). This C_m value is a conservative estimate considering that the extraction process was not fully optimized by this study, which was not necessary since the C_m value achieved was well below typical mitragynine concentrations in kratom (1.3–2.3%). The method is ideal (i) for prioritizing samples for additional testing using other more time-consuming laboratory-based techniques needed to detect and quantify mitragynine and (ii) for field use at international mail facility (IMF) satellite laboratories to help interdict kratom and prevent this dangerous product from reaching the U.S. supply chain.     

Postmortem Changes of Female External Genitalia and Their Importance in Suspected Sexual Abuse

Examination of the female external genitalia to assess for sexual abuse is performed in living individuals, and the interpretation of the findings is based on evidence-based studies. However, in the deceased, no such studies are available, and postmortem changes could present as suspicious findings that can be mistaken for trauma. Patches of discoloration in the hymen were reported previously in one case as hypostasis (i.e., livor and lividity), and based on this finding, it was listed as a finding that is not associated with trauma. This was a retrospective study that was conducted in the Center of Forensic and Legal Medicine in Dammam, Saudi Arabia over a 4-year period. The study included 30 deceased women in whom photographic documentation of their external genitalia was assessed for postmortem changes. The postmortem interval ranged from less than 24 h to more than 100 days, and the ages of these deceased women were in the 20–40 year-old age group. In cases where the hymen, vagina, and/or fossa navicularis were clearly visible, none of these areas showed any hypostatic discoloration. A comparison between antemortem and postmortem appearance of the hymen in one case clearly showed the absence of hypostatic changes in the hymen. In conclusion, any discoloration of the external genitalia that is detected in a female decedent requires serious consideration.     

Recommendations for Maintaining a Dermestid Beetle Colony (Dermestes maculatus) for Processing Human Remains

Dermestid beetles (Dermestes maculatus De Geer 1774) are small carrion insects characterized by a rounded or oval-shaped body and white abdomen with black markings. Given their natural propensity to consume soft tissue throughout various stages of decomposition, biological anthropologists have sought to use dermestids as a forensic processing method in addition to traditional chemical tissue removal techniques. Although useful, most of the existing academic literature regarding the upkeep of dermestid colonies for skeletal remains processing either lack specificity or are outdated. Additionally, nonacademic sources that contain information regarding dermestid maintenance are often disjointed, resulting in a difficulty to replicate habitat construction and ideal environmental conditions. Therefore, this technical note presents recommendations for anthropologists interested in establishing and maintaining a D. maculatus population. These recommendations are based on our experiences using several dermestid colonies to process five unembalmed human heads, procured from an anatomical gift company for a larger study on gunshot trauma. Aspects of the dermestids’ environment that are crucial for the management of a healthy colony include the type of bedding, food, water, and containment method used, in addition to maintaining appropriate temperature ranges (24–27°C) and humidity levels (35–73%). Although habitat construction and dermestid maintenance involve materials that are relatively inexpensive and readily available, setting up and maintaining a D. maculatus colony can be laborious and time consuming and should only be undertaken when the volume of casework is such that this investment would be offset.     

Pitfalls of Computed Tomography 3D Reconstruction Models in Cranial Nonmetric Analysis*

Many studies in the literature have highlighted the utility of virtual 3D databanks as a substitute for real skeletal collections and the important application of radiological records in personal identification. However, none have investigated the accuracy of virtual material compared to skeletal remains in nonmetric variant analysis using 3D models. The present study investigates the accuracy of 20 computed tomography (CT) 3D reconstruction models compared to the real crania, focusing on the quality of the reproduction of the real crania and the possibility to detect 29 dental/cranial morphological variations in 3D images. An interobserver analysis was performed to evaluate trait identification, number, position, and shape. Results demonstrate a false bone loss in 3D models in some cranial regions, specifically the maxillary and occipital bones in 85% and 20% of the samples. Additional analyses revealed several difficulties in the detection of cranial nonmetric traits in 3D models, resulting in incorrect identification in circa 70% of the traits. In particular, pitfalls included the detection of erroneous position, error in presence/absence rates, in number, and in shape. The lowest percentages of correct evaluations were found in traits localized in the lateral side of the cranium and for the infraorbital suture, mastoid foramen, and crenulation. The present study highlights important pitfalls in CT scan when compared with the real crania for nonmetric analysis. This may have crucial consequences in cases where 3D databanks are used as a source of reference population data for nonmetric traits and pathologies and during bone-CT comparisons for identification purposes.