A novel method for sorting and reassociating commingled human remains using deviation analysis

This study provides an innovative and novel method for sorting commingled human remains at the sacroiliac joint using deviation analyses. Virtual models were created at the University of Tennessee-Knoxville Donated Skeletal Collection from 69 os coxae and 66 sacra using an EinScan-Pro 2× + Handheld Surface Scanner. The shape of the auricular surfaces was analyzed in Geomagic Wrap 2017, and the congruency of the two auricular surfaces was measured using a deviation analysis. ROC curves were performed on a reference sample composed of 200 commingled and non-commingled joint pairs to identify threshold values that could help sort the commingled remains. A validation sample of 225 pairs was subsequently analyzed to demonstrate the efficacy of this new method on a sample of unknown individuals. Statistical analyses demonstrated that the deviation analysis values from sacroiliac joints of commingled pairs were significantly larger than those from non-commingled individuals (p < 0.0001). Based on the selected threshold values, 98%–100% of pairs were correctly sorted and reassociated. This novel and objective technique improves upon previously subjective strategies for sorting commingled remains and, in the future, will be applied to additional joint surfaces.     

On the challenge of unambiguous identification of fentanyl analogs: Exploring measurement diversity using standard reference mass spectral libraries

Fentanyl analogs are a class of designer drugs that are particularly challenging to unambiguously identify due to the mass spectral and retention time similarities of unique compounds. In this paper, we use agglomerative hierarchical clustering to explore the measurement diversity of fentanyl analogs and better understand the challenge of unambiguous identifications using analytical techniques traditionally available to drug chemists. We consider four measurements in particular: gas chromatography retention indices, electron ionization mass spectra, electrospray ionization tandem mass spectra, and direct analysis in real time mass spectra. Our analysis demonstrates how simultaneously considering data from multiple measurement techniques increases the observable measurement diversity of fentanyl analogs, which can reduce identification ambiguity. This paper further supports the use of multiple analytical techniques to identify fentanyl analogs (among other substances), as is recommended by the Scientific Working Group for the Analysis of Seized Drugs (SWGDRUG).     

The effectiveness of various strategies to improve DNA analysis of formaldehyde-damaged tissues from embalmed cadavers for human identification purposes

Formalin-fixed tissues provide the medical and forensic communities with alternative and often last resort sources of DNA for identification or diagnostic purposes. The DNA in these samples can be highly degraded and chemically damaged, making downstream genotyping using short tandem repeats (STRs) challenging. Therefore, the use of alternative genetic markers, methods that pre-amplify the low amount of good quality DNA present, or methods that repair the damaged DNA template may provide more probative genetic information. This study investigated whether whole genome amplification (WGA) and DNA repair could improve STR typing of formaldehyde-damaged (FD) tissues from embalmed cadavers. Additionally, comparative genotyping success using bi-allelic markers, including INDELs and SNPs, was explored. Calculated random match probabilities (RMPs) using traditional STRs, INDEL markers, and two next generation sequencing (NGS) panels were compared across all samples. Overall, results showed that neither WGA nor DNA repair substantially improved STR success rates from formalin-fixed tissue samples. However, when DNA from FD samples was genotyped using INDEL and SNP-based panels, the RMP of each sample was markedly lower than the RMPs calculated from partial STR profiles. Therefore, the results of this study suggest that rather than attempting to improve the quantity and quality of severely damaged and degraded DNA prior to STR typing, a more productive approach may be to target smaller amplicons to provide more discriminatory DNA identifications. Furthermore, an NGS panel with less loci may yield better results when examining FD samples, due to more optimized chemistries that result in greater allelic balance and amplicon coverage.     

Toward maintaining canine training aid integrity: Visualizing contamination and testing storage materials for a non-hazardous canine training aid

Contamination of canine training aids is a pervasive issue that may lead to incorrect canine discrimination of target odors. It is therefore important to properly store training materials to maintain their integrity and efficiency. First, this study demonstrated the potential for contamination using GloGerm™ as a proxy for odor/particulate transfer. Then, eight types of containers were evaluated to determine (1) the ability to prevent odor permeation and (2) the likelihood of maintaining the ab/adsorbed odor. Lastly, a longitudinal study evaluated how the permeation of the target odor changed over time. Analysis occurred using a direct analysis in real-time mass spectrometer (DART-MS) to detect triacetone triperoxide (TATP) from the non-hazardous canine training aid known as the polymer odor capture-and-release (POCR) system. Results showed that Mylar and Opsak bags were most effective for short-term storage, maintaining low levels of ab/adsorption. Over time, the amount of TATP permeating through the primary containers and collecting in a secondary container (i.e., outer packaging) increased at 1 week and decreased thereafter (up to 4 months). The amount of TATP collecting in the primary containers, however, increased up to 1 month and decreased thereafter.     

Alternative direct-to-amplification cell lysis techniques for forensically relevant non-sperm cells

While efforts have been made to reduce the pervasive backlog of sexual assault evidence collection kits, the actual laboratory process remains very time-consuming due to the requirement of a differential lysis step before DNA purification, as well as intricate mixture analysis towards the end of the DNA workflow. Recently, an alternative, direct-to-amplification sperm lysis method (using 1 M NaOH) was identified. However, a direct cell lysis method for non-sperm cells has not been identified yet. Thus, the primary objective of this work was to find an alternative method that is quick, inexpensive, and does not require multiple purification steps for the lysis of non-sperm cells in sexual assault samples. In this study, vaginal swab samples were lysed with the control method, prepGEM™, as well as six alternative reagents: alkaline buffer with 25–200 mM NaOH, high-salt stain extraction buffer, modified radioimmunoprecipitation assay (RIPA) buffer, mammalian protein extraction reagent (M-PER™), digitonin buffer, and urea/thiourea buffer. Quantification using Quantifiler® Trio of vaginal and semen lysates revealed that the alkaline (25 mM NaOH) and M-PER™ methods were efficient for the lysis of vaginal epithelial cells without substantial sperm cell lysis. Following quantification, analysis of STR profiles from vaginal lysates revealed that the M-PER™ method showed promising results across all metrics examined, including the percentage of detected STR alleles, mean peak heights, peak height ratio, and interlocus balance. Thus, this method was recommended as an alternative to the traditional differential lysis method for non-sperm cells given its ability to produce amplification-ready lysates without any DNA purification step.     

The effects of certain intermediary target materials and their proximity to ballistic gelatin on jacketed hollow point bullet expansion

Expanding bullets are preferred by law enforcement because of their wounding potential and ability to avoid over-penetration which could result in unintended targets being struck by bullets that perforate their intended targets. Expansion failure for jacketed hollow point (JHP) bullets is commonly attributed to several causes including damage to the bullet's cavity, velocity loss, bullet destabilization and materials from intermediate targets filling the bullet's cavity which can cause expansion failure when the bullet subsequently impacts a soft, fluid-based target such as human tissue or ballistic gelatin. In this study, JHP bullets were fired into ballistic gelatin after passing through selected intermediate targets representing items common to shooting incidents. Velocity loss and bullet destabilization were not factors that contributed to the JHP bullets that experienced expansion failure; however, materials obstructing the bullets' cavities and damage to the bullets' cavities were considered causes for some of the JHP bullet expansion failures. It was determined through this research that most of the target materials caused JHP bullet expansion failure when shored against the ballistic gelatin, but when placed at distances of 7 ft. from the ballistic gelatin, bullets fired through the same target materials did expand. This original and unique study produced findings that are of significant value to shooting incident reconstruction experts and other forensic professionals as shooting incidents can call into question a victim's proximity to a wall or door when a bullet(s) perforated such a target material.     

Field-portable detection of fentanyl and its analogs: A review

The need to detect fentanyl and its analogs in the field is an important capability to help prevent unintentional exposure or overdose on these substances, which may result in death. Many portable methods historically used in the field by first responders and other field users to detect and identify other chemical substances, such as hazardous materials, have been applied to the detection and identification of these synthetic opioids. This paper describes field portable spectroscopic methods used for the detection and identification of fentanyl and its analogs. The methods described are automated colorimetric tests including lateral flow assays; vibrational spectroscopy (mid-infrared and Raman); gas chromatography–mass spectrometry; ion mobility spectrometry, and high-pressure mass spectrometry. In each case the background and key details of these technologies are outlined, followed by a discussion of the application of the technology in the field. Attention is paid to the analysis of complex mixtures and limits of detection, including the required spectral databases and algorithms used to interrogate these types of samples. There is also an emphasis on providing actionable information to the (likely) non-scientist operators of these instruments in the field.     

Rapid enantiomeric analysis of zopiclone in serum by supercritical fluid chromatography–tandem mass spectrometry

Zopiclone (ZOP) is a hypnotic drug prescribed to treat insomnia. Due to the chiral nature of ZOP, the psychologically active S-form and inactive R-form need to be determined enantiomerically in a forensic drug analysis. In the present study, a supercritical fluid chromatography (SFC) method was designed with a faster analysis ability than that of previously reported techniques. The SFC–tandem mass spectrometry (SFC–MS/MS) method was optimized using a column with a chiral polysaccharide stationary phase (Trefoil CEL2). ZOP was extracted from pooled human serum using solid-phase extraction (Oasis HLB) and analyzed. The developed SFC–MS/MS method achieved the baseline separation of S-ZOP and R-ZOP within 2 min. The fit-for-purpose method validation indicated that the optimized solid-phase extraction achieved near complete recovery and approximately 70% of the matrix effect. Both the retention time and peak area showed sufficient precision. The lower and upper limits of quantification (LOQ) were 5.7 × 10⁻² ng/mL and 25 ng/mL for R-ZOP, and 5.2 × 10⁻² ng/mL and 25 ng/mL for S-ZOP. The calibration line was linear in the range from lower LOQ to upper LOQ. The stability test indicated that ZOP in serum stored in a refrigerator (4°C) degraded and about 55% remained in 31 days. The quick analysis of the SFC–MS/MS method makes it a valid option for the enantiomeric analysis of ZOP.     

Radiographic age estimation based on degenerative changes of vertebrae

Age estimation is an important component of decedent identification. When assessing adult remains, anthropologists frequently use gross examination of skeletal elements, such as clavicles, ribs, and pubic symphyses. For fleshed bodies, this requires the removal of these elements and maceration prior to analysis. A new method was developed using radiographic imaging to estimate age from degenerative changes of the lower thoracic and upper lumbar vertebrae. This technique will complement anthropological age estimation methods in young and middle-aged adults and may serve as a stand-alone method for older individuals. Digital radiographs from 240 medical examiner cases were evaluated. The sample included 120 females and 120 males between the ages of 18 and 101 years. A 3-phased scoring system was used for the target vertebrae. Transition analysis was conducted on binned average scores and a Bayesian approach was used to assign age intervals. At the 90% credible interval, individuals in Bin 1 were under 36 years of age while those in Bin 3 were over 47 years of age. Individuals in Bin 2 showed too much age variation to be informative. No significant differences were found between males and females. These findings will be especially useful in the age estimation of older adults and may eliminate the need for skeletal sampling in medicolegal cases where advanced degenerative changes are radiographically observed in the lower thoracic and/or upper lumbar vertebrae. This method was developed for use on fleshed individuals but may also be applicable to skeletonized remains.