Potential Use of Bacterial Community Succession in Decaying Human Bone for Estimating Postmortem Interval,,

Bacteria are taphonomic agents of human decomposition, potentially useful for estimating postmortem interval (PMI) in late‐stage decomposition. Bone samples from 12 individuals and three soil samples were analyzed to assess the effects of decomposition and advancing time on bacterial communities. Results indicated that partially skeletonized remains maintained a presence of bacteria associated with the human gut, whereas bacterial composition of dry skeletal remains maintained a community profile similar to soil communities. Variation in the UniFrac distances was significantly greater between groups than within groups (p < 0.001) for the unweighted metric and not the weighted metric. The members of the bacterial communities were more similar within than between decomposition stages. The oligotrophic environment of bone relative to soft tissue and the physical protection of organic substrates may preclude bacterial blooms during the first years of skeletonization. Therefore, community membership (unweighted) may be better for estimating PMI from skeletonized remains than community structure (weighted).     

Development of a PCR High‐Resolution Melt Assay for Artemisia absinthium (Wormwood) and a Triplex Assay with Two Additional “Unregulated Legal High” Species Datura stramonium (Jimson Weed) and Merremia tuberosa (Hawaiian Woodrose),

Artemisia absinthium (wormwood), a common ingredient in absinthe, contains the compound thujone, which is unregulated by the U.S. Drug Enforcement Agency. Thujone can cause an “unregulated legal high” in higher concentrations. The European Union limits thujone from Artemisia species to 35 mg/kg while the U.S. Food and Drug Administration requires less than 10 ppm to be “thujone‐free.” However, individuals can smoke or ingest A. absinthium in different forms. This study developed a polymerase chain reaction (PCR) high‐resolution melt (HRM) assay to detect and identify A. absinthium based on primer specificity, sensitivity, repeatability, and robustness. A triplex assay was performed with three “unregulated legal high” species: Datura stramonium, Merremia tuberosa, and A. absinthium; the PCR HRM assay detected and identified each plant at melt temperatures 77.42 ± 0.20°C, 83.88 ± 0.22°C, and 87.77 ± 0.15°C, respectively. The primer set developed distinguished A. absinthium from a variety of plant species and was successfully triplexed.     

Morphoscopic Trait Expression in “Hispanic” Populations

This study evaluates population variation of eight cranial morphoscopic traits using samples of known southwest Hispanics (n = 72), Guatemalans (n = 106), American Blacks (n = 146), and American Whites (n = 218). We applied the support vector machine (SVM) method to build a prediction model based on a subsample (20%) of the data; the remainder of the data was used as a test sample. The SVM approach effectively differentiated between the four groups with correct classification rates between 72% (Guatemalan group) and 94% (American Black group). However, when the Guatemalan and southwest Hispanic samples were pooled, the same model correctly classified all groups with a higher degree of accuracy (American Black = 96%; American White = 77%; and the pooled Hispanic sample = 91%). This study also identified significant differences between the two Hispanic groups in six of the eight traits using univariate statistical tests. These results speak to the unique population histories of these samples and the current use of the term “Hispanic” within forensic anthropology. Finally, we argue that the SVM can be used as a classification model for ancestry estimation in a forensic context and as a diagnostic tool may broaden the application of morphoscopic trait data for the assessment of ancestry.     

The influence of alcohol content variation in UK packaged beers on the uncertainty of calculations using the Widmark equation

It is common for forensic practitioners to calculate an individual's likely blood alcohol concentration following the consumption of alcoholic beverage(s) for legal purposes, such as in driving under the influence (DUI) cases. It is important in these cases to be able to give the uncertainty of measurement on any calculated result, for this reason uncertainty data for the variables used for any calculation are required. In order to determine the uncertainty associated with the alcohol concentration of beer in the UK the alcohol concentration (%v/v) of 218 packaged beers (112 with an alcohol concentration of ≤5.5%v/v and 106 with an alcohol concentration of >5.5%v/v) were tested using an industry standard near infra-red (NIR) analyser. The range of labelled beer alcohol by volume (ABV's) tested was 3.4%v/v – 14%v/v. The beers were obtained from a range of outlets throughout the UK over a period of 12?months. The root mean square error (RMSE) was found to be ±0.43%v/v (beers with declared %ABV of ≤5.5%v/v) and ±0.53%v/v (beers with declared %ABV of >5.5%v/v) the RMSE for all beers was ±0.48%v/v. The standard deviation from the declared %ABV is larger than those previously utilised for uncertainty calculations and illustrates the importance of appropriate experimental data for use in the determination of uncertainty in forensic calculations.     

Determination of Nitrite in Whole Blood by High‐Performance Liquid Chromatography with Electrochemical Detection and a Case of Nitrite Poisoning

Although nitrite is widely used in meat processing, it is a major toxicity hazard to children and is responsible for the blue‐baby syndrome. A simple and effective method to determine nitrite in whole blood has been devised using ion chromatography with suppressed conductivity detection. The blood sample was deproteinized by adding acetonitrile and purified with mini‐cartridges to remove hydrophobic compounds, chloride ions, and metal ions. An aliquot of the filtrate was injected onto the ion chromatography. The retention time for nitrite was 13.8 min and the detection limit of nitrite in whole blood was 0.4 μmol/L. The calibration curve was linear (r² = 0.9999) over the concentration working range. The blood nitrite concentration of a victim who attempted suicide by ingesting sodium nitrite powder was determined using the present method. The basal levels for nitrite in human blood was determined with 7.1 ± 0.9 μmol/L (n = 12).     

Temperature‐Dependent Postmortem Changes in Human Cardiac Troponin‐T (cTnT): An Approach in Estimation of Time Since Death

Estimation of time of death is an indispensible requirement of every medico‐legal autopsy, but unfortunately, there is not a single method by which it could be determined accurately. This study focused on the temperature‐dependent postmortem degradation of cardiac troponin‐T and its association with postmortem interval (PMI) in human. The analysis involved extraction of the protein, separation by denaturing gel electrophoresis (SDS‐PAGE), and visualization by Western blot using cTnT‐specific monoclonal antibodies. The area of the bands within a lane was quantified by scanning and digitizing the image using Gel Doc (Universal Hood). The results indicate a characteristic banding pattern among human cadavers (n = 6) and a pseudo‐linear relationship between percentage of cTnT degradation and the log of the time since death (r > 0.95), which can be used to estimate the postmortem interval. The data presented demonstrate that this technique can provide an extended time range during which PMI can be more accurately estimated.     

Forensic Autopsies can Determine Latent Prostate Cancer Prevalence

To evaluate the usefulness of forensic autopsies in determining latent prostate cancer (PC) prevalence, we examined latent PC prevalence from autopsies and compared our findings between decedents with and without cancer. Data from forensic autopsies performed in Japan from 2004 to 2014 were obtained. For each prostate, histopathological examinations were performed in both the base and the apex sections. Three hundred and seventeen Japanese decedents were selected for analysis. The mean age of decedents was 56.4 ± 17.8 years (range, 14–94 years). Among this population, 39.4% died suddenly of disease and 60.6% died of external causes. Latent PC was identified in 45 (14.2%) decedents, who ranged from 27 to 93 years old (mean, 71.1 ± 12.9 years). The prevalence of clinically significant PC with a Gleason score of 7 or more was 8.8%, and the rate increased with age. Fifteen males had cancers other than PC. The prevalence of overall latent PC was significantly higher for those with cancer compared with those without (40.0% vs. 12.9%; P = 0.003). In this study, the use of forensic autopsy materials provided the opportunity to obtain a more accurate natural history of PC, as the decedents in this situation would have been more likely to have died suddenly while behaving as normal prior to death, and less likely to have been impacted by long-term medical interventions.     

Forensic Dental Age Estimation by Measuring Root Dentin Translucency Area Using a New Digital Technique

Dentin translucency measurement is an easy yet relatively accurate approach to postmortem age estimation. Translucency area represents a two‐dimensional change and may reflect age variations better than length. Manually measuring area is challenging and this paper proposes a new digital method using commercially available computer hardware and software. Area and length were measured on 100 tooth sections (age range, 19–82 years) of 250 μm thickness. Regression analysis revealed lower standard error of estimate and higher correlation with age for length than for area (R = 0.62 vs. 0.60). However, test of regression formulae on a control sample (n = 33, 21–85 years) showed smaller mean absolute difference (8.3 vs. 8.8 years) and greater frequency of smaller errors (73% vs. 67% age estimates ≤ ±10 years) for area than for length. These suggest that digital area measurements of root translucency may be used as an alternative to length in forensic age estimation.     

Combined DNA Typing and Protein Identification from Unfired Brass Cartridges,,,

Biological evidence analysis from contact traces is adversely affected by low quantity and quality of DNA. Proteins in these samples contain potentially individualizing information and may be particularly important for difficult surfaces such as brass, where DNA may yield incomplete profiles. In this study, touched unfired brass cartridges were sampled using dry tape or wet swabs and analyzed by separating DNA and protein from the same collected material, thus producing both genomic and proteomic information. DNA recovery was similar for both collection methods, with tape yielding an average of 1.36 ± 1.87 ng and swabs, 1.34 ± 3.04 ng. Analysis by mass spectrometry identified 95 proteins, with the two collection methods showing no significant difference (p = 0.76) in the average number of collected proteins: 44.5 ± 10.9, (tape) versus 47.9 ± 20.4 (swabs). Proteins can be collected from fingerprints at levels necessary to provide identifying information, thus expanding information obtained from challenging evidence.     

Age Estimation from Radiographic Images of the Knee

This study examines the reliability of age estimation utilizing the Pyle and Hoerr atlas in relation to a modern Scottish population. The knee radiographs of 442 individuals (168 females, 274 males) were age assessed using the Pyle and Hoerr atlas. Analysis showed that there was a strong correlation between chronological age and estimated age (females R² = 0.968, males R² = 0.952). For females, the atlas method was most accurate between the ages of 9 and 15 years of age with an underage of 2.27 months and an overage of 2.38 months. For males, the atlas consistently overestimated age from the age of 9 years to the age of 16 years from between 0.14 and 8.81 months. The standard deviation for females was 9.86 months and for males was 10.75 months. This study showed that the Pyle and Hoerr atlas can be applied to a modern population with small modifications.     

Relationship Between Self‐Injurious Behaviors and Levels of Aggression in Children and Adolescents Who Were Subject to Medicolegal Examination,

Aggression, which is defined as a behavior causing harm or pain, is a behavioral pattern typically expected in children and adolescents who are involved in criminal activities. The aim of this study was to examine the relationship between aggression and self‐injurious behavior (SIB) in children and adolescents. The study was performed in 295 cases which were sent for medicolegal examination. The mean age of the subjects was 14.27 ± 1.05 years (age range 10–18 years). The aggression levels of the subjects were determined using the Aggression Questionnaire (AQ), which is an updated form of the Buss‐Durkee Hostility Inventory. The mean total AQ score of the subjects with and without SIB was 78.04 ± 21.0 and 62.75 ± 18.05, respectively (p < 0.01). There were significant statistical differences between the two groups with respect to their subscale scores (p < 0.01). It was concluded that the levels of aggression increased in children and adolescents who were involved in criminal activities when the SIBs increased.     

Age Estimation Based on DNA Methylation Using Blood Samples From Deceased Individuals

Age estimation using DNA methylation levels has been widely investigated in recent years because of its potential application in forensic genetics. The main aim of this study was to develop an age predictor model (APM) for blood samples of deceased individuals based in five age-correlated genes. Fifty-one samples were analyzed through the bisulfite polymerase chain reaction (PCR) sequencing method for DNA methylation evaluation in genes ELOVL2, FHL2, EDARADD, PDE4C, and C1orf132. Linear regression was used to analyze relationships between methylation levels and age. The model using the highest age-correlated CpG from each locus revealed a correlation coefficient of 0.888, explaining 76.3% of age variation, with a mean absolute deviation from the chronological age (MAD) of 6.08 years. The model was validated in an independent test set of 19 samples producing a MAD of 8.84 years. The developed APM seems to be informative and could have potential application in forensic analysis.     

Effect of Temperature on Six Different Developmental Landmarks within the Pupal Stage of the Forensically Important Blowfly Calliphora vicina (Robineau‐Desvoidy) (Diptera: Calliphoridae)

This study investigates the pupal development times of the blow fly Calliphora vicina, which were studied in the laboratory at six different constant temperatures (15, 20, 23, 25, 28, and 30°C each ± 1°C). Lower thresholds (t_L) for development were estimated from the linear regression of the developmental rates on each temperature. These data have made it possible to calculate the accumulated degree days (ADD) necessary for C. vicina to complete the larval stage and to achieve adult emergence. The minimal duration of development from oviposition to adult emergence was found to be inversely related to temperature. Additionally, six landmarks in pupal development are showed, and for each of the landmarks, the ADD value was calculated for every rearing temperature involved. These data assist in calculating the duration of the pupal stage based on morphological characteristics and would be of great value for future forensic entomological casework.     

Validity Comparison of Three Dental Methods for Age Estimation Based on Tooth Root Translucency

Age estimation of nonliving subjects plays a very important role in identifying human remains. When adult remains are in question, this process becomes even more complex. Among the numerous dental methods that exist for adults aging, the evaluation of root translucency is one of the most useful. This study studies the correlation between age and root translucency by investigating monoradicular teeth affected by pathological processes using the equations proposed by Lamendin, Prince & Ubelaker and from Singhal et al. The average age values calculated according to the three methods were compared with the values of known age. Age estimation using the Student's t‐test resulted as more accurate in the methods by Singhal et al. and by Prince & Ubelaker, with respect to that of Lamendin. Age estimation based solely on the percentage of radicular dentinal translucency has been found to be more accurate when diseased teeth are involved.     

Reconstruction of Banknote Fragments Based on Keypoint Matching Method

Banknotes may be shredded by a scrap machine, ripped up by hand, or damaged in accidents. This study proposes an image registration method for reconstruction of multiple sheets of banknotes. The proposed method first constructs different scale spaces to identify keypoints in the underlying banknote fragments. Next, the features of those keypoints are extracted to represent their local patterns around keypoints. Then, similarity is computed to find the keypoint pairs between the fragment and the reference banknote. The banknote fragments can determine the coordinate and amend the orientation. Finally, an assembly strategy is proposed to piece multiple sheets of banknote fragments together. Experimental results show that the proposed method causes, on average, a deviation of 0.12457 ± 0.12810° for each fragment while the SIFT method deviates 1.16893 ± 2.35254° on average. The proposed method not only reconstructs the banknotes but also decreases the computing cost. Furthermore, the proposed method can estimate relatively precisely the orientation of the banknote fragments to assemble.     

Elemental Analysis of Variably Contaminated Cremains Using X‐ray Fluorescence Spectrometry

Analyzing and identifying skeletal remains becomes increasingly difficult when remains have been cremated, especially in cases where the cremated material may have been intentionally contaminated with nonskeletal material. This study examined the potential of X‐ray fluorescence spectrometry (XRF) to detect the presence of nonskeletal contaminants in samples of cremains. Eleven samples of cremains were variably combined with concrete mix and analyzed using XRF. Photon counts of elements in each sample were analyzed, and the coefficient of determination (R²) using unweighted linear regression as a function of percent cremains was calculated. Results showed that with changes in the proportion of skeletal material and contaminant, there were significant (R² > 0.90) changes in detected levels of phosphorus, potassium, zinc, aluminum, and sulfur. The use of XRF is concluded to be a valid approach in the identification of the presence of nonskeletal material in potentially contaminated cremains.     

The Accuracy of Handheld Pre‐Arrest Breath Test Instruments as a Predictor of the Evidential Breath Alcohol Test Results

Drunk driving is a serious threat to public safety. All available and appropriate tools for curbing this threat should be employed to their full extent. The handheld pre‐arrest breath test instrument (PBT) is one tool for identifying the alcohol‐impaired driver and enforcing drunk driving legislation. A set of data was evaluated (n = 1779) where the PBT instrument was employed in drunk driving arrests to develop a multivariate predictive model. When maintained and operated by trained personnel, the PBT provides a reasonable estimate of the evidential test result within the relevant forensic range (95% prediction interval:  ± 0.003 g/210 L). ROC analysis shows that a multivariate model for PBT prediction of the evidentiary alcohol concentration above versus below the legal limit of 0.08 g/210 L has excellent performance with an AUC of 0.96. These results would be of value in evidential hearings seeking to admit the PBT results in drunk driving trials.     

Facial Soft Tissue Depth Statistics and Enhanced Point Estimators for Craniofacial Identification: The Debut of the Shorth and the 75‐Shormax

Several methods that have customarily been used in craniofacial identification to describe facial soft tissue depths (FSTDs) implore improvement. They include the calculation of arithmetic means for skewed data, omission of concern for measurement uncertainty, oversight of effect size, and misuse of statistical significance tests (e.g., p‐values for strength of association). This paper redresses these limitations using FSTDs from 10 prior studies (N = 516). Measurement uncertainty was large (>20% of the FSTD), skewness (≥0.8) existed at 11 of the 23 FSTD landmarks examined, and sex and age each explained <4% of the total FSTD variance (η² calculated as part of MANOVA). These results call for a new and improved conceptualization of FSTDs, which is attained by the replacement of arithmetic means with shorths and 75‐shormaxes. The outcomes of this implementation are dramatic reduction in FSTD complexity; improved data accuracy; and new data‐driven standards for casework application of methods.     

Detection and Identification of Kratom (Mitragyna speciosa) and Marijuana (Cannabis sativa) by a Real-Time Polymerase Chain Reaction High-Resolution Melt Duplex Assay,

Mitragyna speciosa (MS), a plant commonly known as kratom, is a widely used “legal high” opiate alternative for pain relief. DNA extracted from MS and 26 additional plant species was amplified by PCR using primers targeting the strictosidine beta-D-glucosidase (SGD) and secologanin synthase 2 (SLS2) genes and detected by high-resolution melt curves using three intercalating dyes. Amplicon sizes were confirmed using agarose gel electrophoresis. The observed melt temperatures for SGD and SLS2 were 77.08 ± 0.38°C and 77.61 ± 0.46°C, respectively, using SYBR^® Green I; 80.18 ± 0.27°C and 80.59 ± 0.08°C, respectively, using Radiant^™ Green; and 82.19 ± 0.04°C and 82.62 ± 0.13°C, respectively, using the LCGreen^® PLUS dye. The SLS2 primers demonstrated higher specificity and identified MS DNA at 0.05 ng/μL. In a duplex reaction, SLS2 and tetrahydrocannabinoic acid synthase gene primers detected and differentiated MS and Cannabis sativa (CS) by melt peaks at 82.63 ± 0.35°C and 85.58 ± 0.23°C, respectively, using LCGreen^® PLUS.     

The Effect of Soft Tissue on Temperature Estimation from Burnt Bone Using Fourier Transform Infrared Spectroscopy

This study investigated the effect of soft tissue and different exposure times on the prediction of burning temperatures of bone when using Fourier transform infrared spectroscopy (FTIR). Ovis aries rib bones were burnt at different temperatures and for varying time intervals. Results of a linear regression analysis indicated that burn temperatures can be predicted with a standard error of ±70°C from defleshed bone spectra. Exposure time does not have a significant impact on prediction accuracy. The presence of soft tissue has a significant impact on heat‐induced changes of the bone matrix in low (<300°C) as well as high temperatures (>800°C), slowing down combustion in the former and accelerating it in the latter (p < 0.05). At medium temperatures, no significant difference was noted. These results provide forensic investigators a new perspective with which to interpret the results of crystallinity measures derived from burnt bone.