Indicator Of Postmortem Interval Research Articles

Bone citrate content as an indicator of post-mortem interval.

The citrate levels in the bone start to decrease after death. Therefore, it may be possible to estimate the post-mortem interval if the citrate content decreases at regular and predictable intervals. To verify this, it was used twelve fresh porcine tibiae and femora, fifteen recently buried porcine bones, two samples of human bones curated from the anatomical teaching collection from Bournemouth University, UK (never been buried) and fifteen samples of archaeological human bones from Saint Augustine the Less, Bristol, UK. The results obtained in this study align together with the data available in the literature, with higher citrate content in the fresh porcine bones and lower in the archaeological samples, however, a statistical significance was not found. Moreover, the formula used to estimate the time since death largely overestimated the known PMI. Further research should be done with a larger sample and a new formula to estimate the post-mortem interval is required.

Unearthing Nature's Cleanup Crew: A Comprehensive Review of Beetle Succession on Vertebrate Corpses

The study of beetles to criminal or legal cases also comes under Forensic entomology. Entomologists have understood for many years that the beetles found on dead bodies as well as the insects convey a story. Decomposition is a complex ecological process in which various organisms, including insects, play critical roles. Among these insects, beetles (Order Coleoptera) are primary decomposers and are integral to the successional process on vertebrate corpses. Beetles frequently occur in the later phases of the decomposition process, which is crucial for the body's dry bones. They are typically discovered on more decomposed corpses, and scientists have been able to calculate the PMI (Post Mortem Interval) by investigating them. Because the variety of beetles on a body changes through time, it can provide as proof of ecological succession. This paper deals with the biology of beetle succession on the corpse. 
 This paper explores the ecological significance of beetle succession on vertebrate corpses, focusing on their taxonomic diversity, life histories, and forensic applications. The paper also discusses the potential for beetles to serve as indicators of post-mortem interval (PMI) in forensic investigations.

In Vitro Animal Model for Estimating the Time since Death with Attention to Early Postmortem Stage.

Estimating the postmortem interval (PMI) has remained the subject of investigations in forensic medicine for many years. Every kind of death results in changes in metabolites in body tissues and fluids due to lack of oxygen, altered circulation, enzymatic reactions, cellular degradation, and cessation of anabolic production of metabolites. Metabolic changes may provide markers determining the time since death, which is challenging in current analytical and observation-based methods. The study includes metabolomics analysis of blood with the use of an animal model to determine the biochemical changes following death. LC-MS is used to fingerprint postmortem porcine blood. Metabolites, significantly changing in blood after death, are selected and identified using univariate statistics. Fifty-one significant metabolites are found to help estimate the time since death in the early postmortem stage. Hypoxanthine, lactic acid, histidine, and lysophosphatidic acids are found as the most promising markers in estimating an early postmortem stage. Selected lysophosphatidylcholines are also found as significantly increased in blood with postmortal time, but their practical utility as PMI indicators can be limited due to a relatively low increasing rate. The findings demonstrate the great potential of LC-MS-based metabolomics in determining the PMI due to sudden death and provide an experimental basis for applying this attitude in investigating various mechanisms of death. As we assume, our study is also one of the first in which the porcine animal model is used to establish PMI metabolomics biomarkers.

Postmortem submersion interval estimation of cadavers recovered from freshwater based on gut microbial community succession.

Microbial community succession during decomposition has been proven to be a useful tool for postmortem interval (PMI) estimation. Numerous studies have shown that the intestinal microbial community presented chronological changes after death and was stable in terrestrial corpses with different causes of death. However, the postmortem pattern of intestinal microbial community succession in cadavers retrieved from water remains unclear. For immersed corpses, the postmortem submersion interval (PMSI) is a useful indicator of PMI. To provide reliable estimates of PMSI in forensic investigations, we investigated the gut microbial community succession of corpses submersed in freshwater and explored its potential application in forensic investigation. In this study, the intestinal microbial community of mouse submersed in freshwater that died of drowning or CO2 asphyxia (i.e., postmortem submersion) were characterized by 16S rDNA amplification and high-throughput sequencing, followed by bioinformatic analyses. The results demonstrated that the chronological changes in intestinal bacterial communities were not different between the drowning and postmortem submersion groups. α-diversity decreased significantly within 14 days of decomposition in both groups, and the β-diversity bacterial community structure ordinated chronologically, inferring the functional pathway and phenotype. To estimate PMSI, a regression model was established by random forest (RF) algorithm based on the succession of postmortem microbiota. Furthermore, 15 genera, including Proteus, Enterococcus, and others, were selected as candidate biomarkers to set up a concise predicted model, which provided a prediction of PMSI [MAE (± SE) = 0.818 (± 0.165) d]. Overall, our present study provides evidence that intestinal microbial community succession would be a valuable marker to estimate the PMSI of corpses submerged in an aquatic habitat.

Dental DNA as an Indicator of Post-Mortem Interval (PMI): A Pilot Research.

Teeth have proven to be a reliable source of DNA for forensic analysis as the pulp is rich in cells and protected from damaging factors and contamination by dental hard tissues. The pilot study aims to evaluate the feasibility of Next-Generation sequencing analysis on dental pulp to detect genetic mutations in DNA caused by post-mortem cell necrosis. We used a 56-gene oncopanel kit on a sample of 17 teeth extracted from living patients. Time of the tooth avulsion was assumed as death of the individual and Post-mortem Interval (PMI) was the time elapse since the DNA extraction and analysis. Days and Accumulated Degree Days (ADD) were assumed as measures of PMI that ranged between 0 to 34 days. Only 38 of the 56 considered genes proved to be affected by mutations (101), thus being of forensic interest. More specifically, 14 mutations occurred only in a specific range of PMIs/ADD; 67 were detected (alone or as clusters of the same gene) at specific PMI/ADD; 22 occurred at every PMI/ADD, except for some specific intervals. Since dental pulp was not targeted by any oncological diseases and all teeth were intact, vital, and from patients with unremarkable medical history, it could be assumed that mutations were due to post-mortem DNA changes induced by pulp death and the increasing time elapse since death. This pilot study found encouraging results in the application of NGS analysis on dental DNA, especially for PMIs of several days for which the traditional tools for PMI estimation have limitations. Further research on a larger sample of PMI and validation research on a larger sample of PMI and validation of the results are indeed necessary.

An optimization-based study of the impact of different parameters on dna degradation

Human remains exposed to various conditions throughout time are frequently utilized for DNA analysis of tissue or bone for identification reasons. The deterioration and loss of DNA in particular environmental settings have previously created a dilemma for investigators. The post-mortem interval (PMI) or time since death is sometimes the most sought-after piece of information in a medical death investigation. Based on the discovery that DNA degradation has a disproportionate effect on the analysis of bigger genetic loci in particular studies, it was postulated that DNA degradation as a result of autolysis or putrefaction might be useful as a possible rate-of-change indicator of PMI. As a sample, goat liver tissue was used. It was incubated in three testing factors, namely pH, salt content, and sugar concentration, and was compared to a control sample that had not been incubated in any of these parameters. The samples were incubated for 24, 48, and 72 hours. An appropriate Lysis Buffer was used to isolate the DNA. Following the isolation of the DNA, quantitative analysis was performed in a UV-Visible Spectrophotometer (for both the control and incubated samples), followed by Agarose Gel Electrophoresis. It was discovered that the data provided by these studies, taken as a whole, show that we can give information relevant for calculating the post-mortem interval, mostly within the first 72 hours following death. Later, it was decided to use Response Surface Methodology (RSM) to optimize the parameters we had picked, with the BBD design model being the favoured choice. This was done with the help of software named, Design-Expert v11. A Fit Summary that showed a linear model fit for the data. ANOVA was used to generate an equation that may be used to quantify the amount of DNA present in a tissue sample that has been exposed to a specific value of pH, salt concentration, and sugar concentration. Finally, a 3-D response surface curve was produced, two factors at a time, to highlight the variance in DNA loss when those two parameters are taken into account.

Review of Synthesiomyia nudiseta (Diptera: Muscidae) as a useful tool in forensic entomology.

Synthesiomyia nudiseta (van der Wulp, 1883) is a synanthropic muscid found in tropical and subtropical zones around the world. The larvae of this species are a secondary agent of myiasis with necrophagous habits and play an important role in forensic entomology, as they are used as an indicator of post-mortem interval. Adults can be considered vectors of etiological agents such as Escherichia coli and Shigella dysenteriae. Due to its ability to adapt to different environmental conditions, its high dispersal capacity (shown by its introduction to Europe), its predatory habits in the last larval stage and the difficulty of identifying it, a very important goal is to update our knowledge about this species. Therefore, the main objective of this paper is to review the identification, geographical distribution and biology of this species in order to provide better support to investigations involving this fly.

Fresh vs. frozen human decomposition – A preliminary investigation of lipid degradation products as biomarkers of post-mortem interval

Traditionally, the post-mortem interval (PMI) is determined using methods such as forensic entomology or forensic pathology, however, these techniques are often limited to a particular post-mortem window of up to 72 – 120 h after death. In this study, lipids extracted from decomposing human tissue were investigated as potential soft-tissue biomarkers of PMI. Tissue samples were collected from two whole human donors (n = 2), one frozen and one fresh, placed at the Australian Facility for Taphonomic Experimental Research (AFTER) over the course of 69 days post-placement. These samples were analysed using gas chromatography coupled with tandem mass spectrometry (GC–MS/MS), demonstrating statistically significant differences for most fatty acid analytes and further highlighting the well-known stability of sterol compounds over time. Differences were observed between the fresh and frozen donor, with the fresh donor displaying more distinct stages of decomposition. Chemical differences between the donors were more distinct in the fatty acids than the sterols. The fatty acid profiles over time were further investigated and target analytes comprising saturated fatty acids (stearic acid and palmitic acid) and unsaturated fatty acids (oleic acid and linoleic acid) were found to be of particular importance due to their potential as indicators of PMI.

Progressive dehydration in decomposing bone: a potential tool for forensic anthropology

The aim of this pilot study was to determine whether collagen and/or water content of bone vary during soft tissue putrefaction by thermogravimetric analysis with a view to eventually developing a possible forensic application to determine post-mortem interval. Porcine bone decomposed in a shallow burial showed an approximate difference in average mass loss of 15 ± 8% when heated between 22 and 100 °C, compared to 14 ± 3% for porcine bone decomposed in a surface deposition, equating to water loss. Mass loss showed peaks at 0, 250–500 and 1200–1500 cumulative cooling degree days’ (CCDD) deposition for the experimental porcine bone. Should these measurements prove consistent in future studies on a wider variety of porcine and eventually human skeletal elements, they may have potential to be corroborated with other data when determining post-mortem interval, especially with disarticulated bones. A downward trend in mass loss was apparent within shallow burial and surface deposition scenarios (inclusive of freeze-dried controls) for the thermolysis of collagen (and other proteins) between 220 and 650 °C during thermogravimetric analysis. This was inconsistent within the time frame examined (0–1450 cumulative cooling degree days), and so demonstrates less potential as an indicator of post-mortem interval during soft tissue putrefaction.

“From womb to tomb; we’re bound to others”: microbiome in forensic science

Microbiome is a new field of interest in clinical medicine with high potential in forensic medicine. It could be used in several applications, such as post-mortem interval (PMI) estimation, personal identification, differential diagnosis of cause of death and toxicology. Regarding PMI, during the decomposition of a corpse, the passage of time involves changing in microbial population both outside and inside the corpse but also in surrounding soil (cadaver decomposition island). These variations could be hypothetically used as PMI indicators (microbial clock), even thanks to the development of machine learning approach. Another potential use of skin and saliva microbiome is personal identification thanks to its inter-individual variability and tendency to remain unvarying over time. It may also be helpful to link a person to a specific object that has been touched (microbial fingerprint). Furthermore, we could infer some information about health state of human subjects, comparing post-mortem and ante-mortem microbiome, but this field of research is quite new and needs further studies. Moreover, we have to consider the influence of microbiome metabolism in post-mortem toxicological evaluation; microbes could alter substances concentrations – for example of ethanol, gamma-hydroxybutyric acid (GHB) and nitrobenzodiazepines – due to enzymatic degradation and individual microbial metabolism. Finally, integration of microbiome and human being’s transcriptomic analysis may be helpful to depict their complex interactions even after death.

Vitamin C as a Potential Indicator of Post-Mortem Interval: A Biochemical Analysis

Accurate estimates of postmortem interval (PMI) are crucial for successful investigation of death especially during un-witnessed death examinations, thus continues to be one of the most strenuous variable to be determined correctly. There are numerous ways to estimate postmortem interval (PMI), or time since death, including body temperature, rigor mortis, insect activity and decomposition. Individually, however, many of these indicators are prone to inaccuracy due to the influence of the external environment upon them. Furthermore, various metabolites have also been identified that change post mortem, but lack of accuracy and reproducibility probably did not let their extensive utilization as a PMI indicator, warranting a simple yet precise marker and easy technique for PMI evaluation. Therefore, present study proposes that vitamin C could be used for precise determination of PMI using simple, easy and rapid biochemical analysis. Vitamin C was estimated at 37°C and 4°C in liver, spleen, kidneys and blood over a time period of 24 hours. Results revealed a gradual change in ascorbic acid levels in the spleen, liver, kidneys and blood from time of death till 24 hours after death indicating its potential to accurately measure time since death.

Characterizing forensically important insect and microbial community colonization patterns in buried remains

During violent criminal actions in which the perpetrator disposes of the victim’s remains by burial, the analysis of insects and bacterial colonization patterns could be necessary for postmortem interval (PMI) estimation. Our research aimed to assess the decomposition process of buried rat carcasses from shallow graves (40 cm), the diversity and dynamics of insects and bacteria throughout the decomposition stages, and the environmental parameters’ influence on these variations. The results provide further insight on decomposition in soil and contribute to a broader understanding of the factors involved in decomposition by qualitatively and quantitatively analysing the decomposer community (bacteria and insects). Additionally, two bacterial taxa, Enterococcus faecalis and Clostridium paraputrificum that were investigated for the first time as PMI indicators using quantitative polymerase chain reaction (qPCR) showed differential abundance over time, promising data for PMI estimation. The current study on the decomposition of buried rat carcasses in a natural environment will strengthen the current knowledge on decomposed remains from shallow graves and represents an effort to quantify insect and bacterial taxa as PMI estimators.

Microbiome Data Accurately Predicts the Postmortem Interval Using Random Forest Regression Models.

Death investigations often include an effort to establish the postmortem interval (PMI) in cases in which the time of death is uncertain. The postmortem interval can lead to the identification of the deceased and the validation of witness statements and suspect alibis. Recent research has demonstrated that microbes provide an accurate clock that starts at death and relies on ecological change in the microbial communities that normally inhabit a body and its surrounding environment. Here, we explore how to build the most robust Random Forest regression models for prediction of PMI by testing models built on different sample types (gravesoil, skin of the torso, skin of the head), gene markers (16S ribosomal RNA (rRNA), 18S rRNA, internal transcribed spacer regions (ITS)), and taxonomic levels (sequence variants, species, genus, etc.). We also tested whether particular suites of indicator microbes were informative across different datasets. Generally, results indicate that the most accurate models for predicting PMI were built using gravesoil and skin data using the 16S rRNA genetic marker at the taxonomic level of phyla. Additionally, several phyla consistently contributed highly to model accuracy and may be candidate indicators of PMI.

Histological transformations of the dental pulp as possible indicator of post mortem interval: a pilot study

BackgroundThe correct estimation of the post mortem interval (PMI) can be crucial on the success of a forensic investigation. Diverse methods have been used to estimate PMI, considering physical changes that occur after death, such as mortis algor, livor mortis, among others. Degradation after death of dental pulp is a complex process that has not yet been studied thoroughly. It has been described that pulp RNA degradation could be an indicator of PMI, however this study is limited to 6 days. The tooth is the hardest organ of the human body, and within is confined dental pulp. The pulp morphology is defined as a lax conjunctive tissue with great sensory innervation, abundant microcirculation and great presence of groups of cell types. AimThe aim of this study is to describe the potential use of pulp post mortem alterations to estimate PMI, using a new methodology that will allow obtainment of pulp tissue to be used for histomorphological analysis. The current study will identify potential histological indicators in dental pulp tissue to estimate PMI in time intervals of 24h, 1 month, 3 months and 6 months. Materials and methodThis study used 26 teeth from individuals with known PMI of 24h, 1 month, 3 months or 6 months. All samples were manipulated with the new methodology (Carrasco, P. and Inostroza C. inventors; Universidad de los Andes, assignee. Forensic identification, post mortem interval estimation and cause of death determination by recovery of dental tissue. United State patent US 61/826,558 23.05.2013) to extract pulp tissue without the destruction of the tooth. The dental pulp tissues obtained were fixed in formalin for the subsequent generation of histological sections, stained with Hematoxylin Eosin and Masson's Trichrome. All sections were observed under an optical microscope using magnifications of 10× and 40×. ResultsThe microscopic analysis of the samples showed a progressive transformation of the cellular components and fibers of dental pulp along PMI. These results allowed creating a chart of qualitative and quantitative parameters to be used on the estimation on PMI based on microscopic degradation of dental pulp. ConclusionsThe histological transformations of dental pulp as a function of time can be used as PMI indicators.

Sex- and Size-Related Patterns of Carrion Visitation in Necrodes littoralis (Coleoptera: Silphidae) and Creophilus maxillosus (Coleoptera: Staphylinidae).

The estimation of postmortem interval (PMI) based on successional patterns of adult insects is largely limited, due to the lack of potential PMI markers. Sex and size of adult insects could be easily used for such estimation. In this study, sex- and size-related patterns of carrion attendance by adult insects were analyzed in Necrodes littoralis (Coleoptera: Silphidae) and Creophilus maxillosus (Coleoptera: Staphylinidae). For both species, abundance of males and females changed similarly during decomposition. A slightly female-biased sex ratio was recorded in N. littoralis. Females of N. littoralis started visiting carcasses, on average, one day earlier than males. There was a rise in size of males of N. littoralis at the end of decomposition, whereas for females of both species and males of C. maxillosus, no size-related patterns of carrion visitation were found. Current results demonstrate that size and sex of adult carrion beetles are poor indicators of PMI.

Changes in Vickers hardness during the decomposition of bone: Possibilities for forensic anthropology

The purpose of this study was to determine how the Vickers hardness (HV) of bone varies during soft tissue putrefaction. This has possible forensic applications, notably for determining the postmortem interval. Experimental porcine bone samples were decomposed in surface and burial deposition scenarios over a period of 6 months. Although the Vickers hardness varied widely, it was found that when transverse axial hardness was subtracted from longitudinal axial hardness, the difference showed correlations with three distinct phases of soft tissue putrefaction. The ratio of transverse axial hardness to longitudinal axial hardness showed a similar correlation. A difference of 10 or greater in HV with soft tissue present and signs of minimal decomposition, was associated with a decomposition period of 250 cumulative cooling degree days or less. A difference of 10 (+/- standard error of mean at a 95% confidence interval) or greater in HV associated with marked decomposition indicated a decomposition period of 1450 cumulative cooling degree days or more. A difference of −7 to +8 (+/- standard error of mean at a 95% confidence interval) was thus associated with 250 to 1450 cumulative cooling degree days’ decomposition. The ratio of transverse axial HV to longitudinal HV, ranging from 2.42 to 1.54, is a more reliable indicator in this context and is preferable to using negative integers These differences may have potential as an indicator of postmortem interval and thus the time of body deposition in the forensic context.

Degradation of Kidney and Psoas Muscle Proteins as Indicators of Post-Mortem Interval in a Rat Model, with Use of Lateral Flow Technology

We investigated potential protein markers of post-mortem interval (PMI) using rat kidney and psoas muscle. Tissue samples were taken at 12 h intervals for up to 96 h after death by suffocation. Expression levels of eight soluble proteins were analyzed by Western blotting. Degradation patterns of selected proteins were clearly divided into three groups: short-term, mid-term, and long-term PMI markers based on the half maximum intensity of intact protein expression. In kidney, glycogen synthase (GS) and glycogen synthase kinase-3β were degraded completely within 48 h making them short-term PMI markers. AMP-activated protein kinase α, caspase 3 and GS were short-term PMI markers in psoas muscle. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was a mid-term PMI marker in both tissues. Expression levels of the typical long-term PMI markers, p53 and β-catenin, were constant for at least 96 h post-mortem in both tissues. The degradation patterns of GS and caspase-3 were verified by immunohistochemistry in both tissues. GAPDH was chosen as a test PMI protein to perform a lateral flow assay (LFA). The presence of recombinant GAPDH was clearly detected in LFA and quantified in a concentration-dependent manner. These results suggest that LFA might be used to estimate PMI at a crime scene.

Evaluation of post-mortem lateral cerebral ventricle changes using sequential scans during post-mortem computed tomography.

In the present study, we evaluated post-mortem lateral cerebral ventricle (LCV) changes using computed tomography (CT). Subsequent periodical CT scans termed “sequential scans” were obtained for three cadavers. The first scan was performed immediately after the body was transferred from the emergency room to the institute of legal medicine. Sequential scans were obtained and evaluated for 24 h at maximum. The time of death had been determined in the emergency room. The sequential scans enabled us to observe periodical post-mortem changes in CT images. The series of continuous LCV images obtained up to 24 h (two cases)/16 h (1 case) after death was evaluated. The average Hounsfield units (HU) within the LCVs progressively increased, and LCV volume progressively decreased over time. The HU in the cerebrospinal fluid (CSF) increased at an individual rate proportional to the post-mortem interval (PMI). Thus, an early longitudinal radiodensity change in the CSF could be potential indicator of post-mortem interval (PMI). Sequential imaging scans reveal post-mortem changes in the CSF space which may reflect post-mortem brain alterations. Further studies are needed to evaluate the proposed CSF change markers in correlation with other validated PMI indicators.Electronic supplementary materialThe online version of this article (doi:10.1007/s00414-016-1327-2) contains supplementary material, which is available to authorized users.

Technical Note: “Mitochondrial and nuclear DNA approaches for reliable identification of Lucilia (Diptera, Calliphoridae) species of forensic interest from Southern Europe”

In forensic entomology, rapid and unambiguous identification of blowfly species is a critical prerequisite for accurately estimating the post-mortem interval (PMI). The conventional diagnosis of cadaveric entomofauna based on external characters is hampered by the morphological similarities between species, especially in immature stages. Genetic analysis has been shown to allow precise and reliable diagnosis and delimitation of insect species. Nevertheless, the taxonomy of some species remains unresolved. This study was focused on improving the effectiveness and accuracy of analysis based on the widely used cytochrome c oxidase subunit I barcode region (COI barcode, 658bp), complemented by other mitochondrial and nuclear regions, such as cytochrome b (Cyt-b, 307bp) and the second internal transcribed spacer (ITS2, 310-331bp), for the identification of Southern European blowflies. We analyzed a total of 209 specimens, collected from 38 human corpses, belonging to three Calliphoridae genera and seven species: Chrysomya (Ch. albiceps), Calliphora (C. vicina and C. vomitoria), and Lucilia (L. sericata, L. ampullacea, L. caesar and L. illustris). These species are the most common PMI indicators in Portugal. The results revealed that unambiguous separation of species of the Lucilia genus requires different loci from the barcode region. Furthermore, we conclude that the ITS2 (310-331bp) molecular marker is a promising diagnostic tool because its inter-specific discriminatory power enables unequivocal and consistent distinctions to be made, even between closely related species (L. caesar-L. illustris). This work also contributes new genetic data that may be of interest in performing species diagnosis for Southern European blowflies. Notably, to the best of our knowledge, we provide the first records of the Cyt-b (307bp) locus for L. illustris and the ITS2 (310-331bp) region for Iberian Peninsula Lucilia species.

Estimating Time Since Death from Postmortem Human Gut Microbial Communities

Postmortem succession of human-associated microbial communities ("human microbiome") has been suggested as a possible method for estimating postmortem interval (PMI) for forensic analyses. Here we evaluate human gut bacterial populations to determine quantifiable, time-dependent changes postmortem. Gut microflora were repeatedly sampled from the proximal large intestine of 12 deceased human individuals as they decayed under environmental conditions. Three intestinal bacterial genera were quantified by quantitative PCR (qPCR) using group-specific primers targeting 16S rRNA genes. Bacteroides and Lactobacillus relative abundances declined exponentially with increasing PMI at rates of Nt=0.977e(-0.0144t) (r2=0.537, p<0.001) and Nt=0.019e(-0.0087t) (r2=0.396, p<0.001), respectively, where Nt is relative abundance at time (t) in cumulative degree hours. Bifidobacterium relative abundances did not change significantly: Nt=0.003e(-0.002t) (r2=0.033, p=0.284). Therefore, Bacteroides and Lactobacillus abundances could be used as quantitative indicators of PMI.