The Brain and Body Donation Program (BBDP) at Banner Sun Health Research Institute (http://www.brainandbodydonationprogram.org) started in 1987 with brain‐only donations and currently has banked more ...than 1600 brains. More than 430 whole‐body donations have been received since this service was commenced in 2005. The collective academic output of the BBDP is now described as the Arizona Study of Aging and Neurodegenerative Disorders (AZSAND). Most BBDP subjects are enrolled as cognitively normal volunteers residing in the retirement communities of metropolitan Phoenix, Arizona. Specific recruitment efforts are also directed at subjects with Alzheimer's disease, Parkinson's disease and cancer. The median age at death is 82. Subjects receive standardized general medical, neurological, neuropsychological and movement disorders assessments during life and more than 90% receive full pathological examinations by medically licensed pathologists after death. The Program has been funded through a combination of internal, federal and state of Arizona grants as well as user fees and pharmaceutical industry collaborations. Subsets of the Program are utilized by the US National Institute on Aging Arizona Alzheimer's Disease Core Center and the US National Institute of Neurological Disorders and Stroke National Brain and Tissue Resource for Parkinson's Disease and Related Disorders. Substantial funding has also been received from the Michael J. Fox Foundation for Parkinson's Research. The Program has made rapid autopsy a priority, with a 3.0‐hour median post‐mortem interval for the entire collection. The median RNA Integrity Number (RIN) for frozen brain and body tissue is 8.9 and 7.4, respectively. More than 2500 tissue requests have been served and currently about 200 are served annually. These requests have been made by more than 400 investigators located in 32 US states and 15 countries. Tissue from the BBDP has contributed to more than 350 publications and more than 200 grant‐funded projects.
Extracellular vesicles (EVs) are involved in a wide range of physiological and pathological processes by shuttling material out of and between cells. Tissue EVs may thus lend insights into disease ...mechanisms and also betray disease when released into easily accessed biological fluids. Since brain-derived EVs (bdEVs) and their cargo may serve as biomarkers of neurodegenerative diseases, we evaluated modifications to a published, rigorous protocol for separation of EVs from brain tissue and studied effects of processing variables on quantitative and qualitative outcomes. To this end, size exclusion chromatography (SEC) and sucrose density gradient ultracentrifugation were compared as final separation steps in protocols involving stepped ultracentrifugation. bdEVs were separated from brain tissues of human, macaque, and mouse. Effects of tissue perfusion and a model of post-mortem interval (PMI) before final bdEV separation were probed. MISEV2018-compliant EV characterization was performed, and both small RNA and protein profiling were done. We conclude that the modified, SEC-employing protocol achieves EV separation efficiency roughly similar to a protocol using gradient density ultracentrifugation, while decreasing operator time and, potentially, variability. The protocol appears to yield bdEVs of higher purity for human tissues compared with those of macaque and, especially, mouse, suggesting opportunities for optimization. Where possible, perfusion should be performed in animal models. The interval between death/tissue storage/processing and final bdEV separation can also affect bdEV populations and composition and should thus be recorded for rigorous reporting. Finally, different populations of EVs obtained through the modified method reported herein display characteristic RNA and protein content that hint at biomarker potential. To conclude, this study finds that the automatable and increasingly employed technique of SEC can be applied to tissue EV separation, and also reveals more about the importance of species-specific and technical considerations when working with tissue EVs. These results are expected to enhance the use of bdEVs in revealing and understanding brain disease.
Researchers have observed that rainfall may re‐initiate decomposition in desiccated tissue; however, no conclusive research‐based evidence exists on the specific effects of rehydration on ...decomposition. Therefore, this study aimed to assess the effects of artificial rehydration on the progression of decomposition following the advanced stage of decomposition. Twelve adult pig cadavers (8 experimental; 4 controls) were placed in the central Highveld of South Africa during cooler (April–July 2021) and warmer (August–November 2021) months. Decomposition was scored approximately biweekly to obtain the total body score, and accumulated degree days (ADD) were calculated for each pig. All pig cadavers were covered by chicken wire cages with transparent tarps to control for natural rehydration and scavenging. Once the experimental pig cadavers reached a three‐visit stasis in the advanced phase of decomposition, they were artificially rehydrated, and changes in the progression of decomposition between the control and experimental groups were plotted (ADD against TBS) for observation. The rehydrated experimental pig cadavers showed re‐initiation of decay and insect re‐colonization, while the control cadavers mainly remained in a state of stasis with insect activity ceased altogether. Greater cadaver decomposition islands and a color change post‐rehydration were also noted in some experimental cadavers. This supports the need for future research on the impact of rehydration, including associated soil moisture on decomposition rates, progression, and invertebrate colonization, which will enhance our understanding of the effects these environmental factors have on the accuracy of post‐mortem interval estimation.
Forensic taphonomic studies are regionally specific and improve time since death estimates for medico‐legal casework. Within forensic taphonomy and carrion ecology, vertebrate scavengers are ...under‐researched with many studies conducted using multiple, unclothed carcasses. This is a forensically unrealistic experimental design choice with unknown impact. The effect of variation in carrion biomass on the decomposition ecosystem, particularly where vertebrate scavengers are concerned, requires clarification. To assess the effect of carrion biomass load on vertebrate scavenging and decomposition rate, seasonal baseline data for single, clothed ~60 kg porcine carcasses were compared to clothed multiple‐carcass deployments, in a forensically relevant habitat of Cape Town, South Africa. Decomposition was tracked via weight loss and bloat progression and scavenging activity via motion‐activated cameras. The single carcasses decayed more quickly, particularly during the cooler, wetter winter, strongly correlated with concentrated Cape gray mongoose (Galerella pulverulenta) scavenging activity. On average and across seasons, the single carcasses lost 68% of their mass by day 32 (567 accumulated degree days ADD), compared to 80 days (1477 ADD) for multi‐carcass deployments. The single carcasses experienced substantially more scavenging activity, with longer visits by single and multiple mongooses, totaling 53 h on average compared to 20 h for the multi‐carcass deployments. These differences in scavenging activity and decay rate demonstrate the impact of carrion biomass load on decomposition for forensic taphonomy research. These findings need corroboration. However, forensic realism requires consideration in taphonomic study design. Longitudinally examining many single carcasses may produce more forensically accurate, locally appropriate, and usable results.
Recent advancements in omics techniques have revolutionised the study of biological systems, enabling the generation of high‐throughput biomolecular data. These innovations have found diverse ...applications, ranging from personalised medicine to forensic sciences. While the investigation of multiple aspects of cells, tissues or entire organisms through the integration of various omics approaches (such as genomics, epigenomics, metagenomics, transcriptomics, proteomics and metabolomics) has already been established in fields like biomedicine and cancer biology, its full potential in forensic sciences remains only partially explored. In this review, we have presented a comprehensive overview of state‐of‐the‐art analytical platforms employed in omics research, with specific emphasis on their application in the forensic field for the identification of the cadaver and the cause of death. Moreover, we have conducted a critical analysis of the computational integration of omics approaches, and highlighted the latest advancements in employing multi‐omics techniques for forensic investigations.
Aims
Decomposition, a complicated process, depends on several factors, including carrion insects, bacteria and the environment. However, the composition of and variation in oral bacteria over long ...periods of decomposition remain unclear. The current study aims to illustrate the composition of oral bacteria and construct an informative model for estimating the post‐mortem interval (PMI) during decomposition.
Methods and Results
Samples were collected from rats' oral cavities for 59 days, and 12 time points in the PMI were selected to detect bacterial community structure by sequencing the V3–V4 region of the bacterial 16S ribosomal RNA (16S rRNA) gene on the Ion S5 XL platform. The results indicated that microorganisms in the oral cavity underwent great changes during decomposition, with a tendency for variation to first decrease and then increase at day 24. Additionally, to predict the PMI, an informative model was established using the random forest algorithm. Three genera of bacteria (Atopostipes, Facklamia and Cerasibacillus) were linearly correlated at all 12 time points in the 59‐day period. Planococcaceae was selected as the best feature for the last 6 time points. The R2 of the model reached 93.94%, which suggested high predictive accuracy. Furthermore, to predict the functions of the oral microbiota, PICRUSt results showed that energy metabolism was increased on day 3 post‐mortem and carbohydrate metabolism surged significantly on days 3 and 24 post‐mortem.
Conclusions
Overall, our results suggested that post‐mortem oral microbial community data can serve as a forensic resource to estimate the PMI over long time periods.
Significance and Impact of the Study
The results of the present study are beneficial for estimating the PMI. Identifying changes in the bacterial community is of great significance for further understanding the applicability of oral flora in forensic medicine.
Dermestes frischii Kugelann, 1792 and Dermestes undulatus Brahm, 1790 are the most abundant species worldwide at outdoor or indoor crime scenes during the dry and skeletal stages of decomposition. ...The attribution of larval age in these beetles is problematic due to the variable number of instars, which is influenced by environmental factors. In this study, a morphometric approach was used to look for potential morphological features as evidence of larval stages. Breeding and monitoring were performed for both species in an incubator with a preset temperature of 28°C ± 0.5 without a photoperiod. Morphometric measurements were made on 10 larvae per instar for each species using length, width, and thickness parameters. Linear discriminant analysis was then used to generate decision boundaries that clearly separated larval stages. The cross‐validation procedure demonstrated that the morphometric approach successfully discriminated adjacent larval stages in both species with high values of sensitivity and specificity. This less‐invasive approach could improve the ability to estimate minPMI in forensic studies of Dermestidae beetles. Future studies may extend this approach to other species and establish good practices for collecting and storing specimens for morphometric analysis.
To date, there have been no studies on necrophagous fly populations on Hainan Island in China. Thus, we investigated the species composition of necrophagous flies as well as their geographic ...distribution on Hainan Island for the first time. Ten sites in different climatic regions across the island were sampled for 7 days per location from November 1, 2018 to February 28, 2019 and from July 1, 2019 to October 31, 2019. Bottle traps made of 1.5 L soft plastic bottles were used to trap necrophagous flies. The collected individuals were identified to species. The specimens represented 5 families and 28 species. Chrysomya megacephala (Fabricius, 1794) was the most dominant species, followed by Chrysomya rufifacies (Macquart, 1843), Hemipyrellia ligurriens (Wiedemann, 1830), Boettcherisca peregrine (Robineau‐Desvoidy, 1830), Parasarcophaga dux (Thomson, 1868), Parasarcophaga misera (Walker, 1849), Synthesiomyia nudiseta (Wulp, 1883), and Ophyra chalcogaster (Wiedemann, 1924). The largest collection of flies was obtained in the semiarid region. Species richness was highest in the subhumid region and was higher in summer than in winter, but there were exceptions, such as L. hainanensis (Fan,1965), Boettcherisca formosensis (Lopes, 1961), and Muscina stabulans (Fallen, 1817). Fannia pusio (Wiedemann, 1830), and Boettcherisca formosensis (Lopes, 1961) were newly recorded species on Hainan Island. Of the necrophagous flies collected during the study, we propose several predominant species based on the criteria of distribution, occurrence frequency, and resource preference. Our results not only investigate necrophagous flies on Hainan Island but also accumulate data for criminal investigations in the region.
Estimates of post-mortem interval (PMI), which often serve as pivotal evidence in forensic contexts, are fundamentally based on assessments of variability among diverse molecular markers (including ...proteins and metabolites), their correlations, and their temporal changes in post-mortem organisms. Nevertheless, the present approach to estimating the PMI is not comprehensive and exhibits poor performance. We developed an innovative approach that integrates multi-omics and artificial intelligence, using multimolecular, multimarker, and multidimensional information to accurately describe the intricate biological processes that occur after death, ultimately enabling inference of the PMI. Called the multi-omics stacking model (MOSM), it combines metabolomics, protein microarray electrophoresis, and fourier transform-infrared spectroscopy data. It shows improved prediction accuracy of the PMI, which is urgently needed in the forensic field. It achieved an accuracy of 0.93, generalized area under the receiver operating characteristic curve of 0.98, and minimum mean absolute error of 0.07. The MOSM integration framework not only considers multiple markers but also incorporates machine-learning models with distinct algorithmic principles. The diversity of biological mechanisms and algorithmic models further ensures the generalizability and robustness of PMI estimation.
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•A comprehensive framework was constructed to analyze the molecular markers detected from three omics.•The MOSM showed superior PMI estimation by effectively integrating diverse molecular markers’ change patterns.•The multi-omics procedure could be a novel tool for forensic and biomedical science.
Estimating the post-mortem interval (PMI) is an essential step in forensic investigations, particularly those involving homicides and unwitnessed deaths. However, traditional methods occasionally ...yield inconsistent estimates. Histological and molecular techniques are considered crucial in forensic pathology and are frequently employed to estimate the time interval of death. The gingiva is an oral mucosal tissue used to estimate PMI. This review aimed to examine the potential of histological methods to determine PMI using oral mucosal tissue, namely the gingiva, and to investigate changes that occur in oral mucosal tissue at different time intervals when compared with those in normal tissues. The oral mucosa comprises layers of stratified squamous epithelium and connective tissue. Similar to other body tissues, changes are known to occur in the gingiva after death, and these cellular and tissue changes should also be considered. Alterations in the gingiva include homogenisation, karyorrhexis, pyknosis, karyolysis, chromatin clumping, eosinophilia, collagen fibre degradation, and the loss of tissue architecture. Reviews collating the results of original trials have consistently reported how the oral mucosa is altered by autolysis and how such changes can be observed in histological tissue morphology after death. Histology is an acceptably accurate technique for estimating PMIs.
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