Postmortem Interval Research Articles

A Biochemical and Histological Assessment of Postmortem Changes to the Eyes of Domestic Pigs: A Preliminary Study

The Postmortem Interval (PMI) is the time from the death of an animal to its discovery. From a veterinary forensic standpoint, an accurate estimation of the PMI is of particular importance, especially with the observed increase in deaths of domestic and wild animals. A preliminary study was conducted using the eyes of domestic pigs. A biochemical analysis was conducted on the vitreous humor of the eye, whilst a histological analysis was conducted on the retina. The eyes were stored at +4 °C and changes were assessed at time intervals of 0, 12, 24, 48, and 120 h. The biochemical analysis during the PMI established a decrease in sodium, chlorine, and glucose concentrations, and a rise in potassium concentration. Accordingly, a simple linear regression showed a significant correlation between changes in concentrations of sodium (Na+), potassium (K+), chloride (Cl−), and glucose, in relation to the PMI. The histological analysis showed evident morphological changes in the retina, which included homogenization of the rod and cone cells, pyknosis of the outer nuclear layer, homogenization of the outer plexiform layer, pyknosis of the inner nuclear layer, homogenization of the inner plexiform layer, and pyknosis of the nuclei of the ganglion layer of the retina.

An effective method for preparing histological sections of blow fly intra-puparial stages for minimum PMI estimations.

Blow flies (Diptera: Calliphoridae) are generally early colonisers of fresh cadavers, enabling the estimation of a minimum post-mortem interval (minPMI) based on an accurate aging of the oldest immature stages associated with a cadaver. In blow flies, the pupal stage and the subsequent development of the adult take place inside a protective case, the puparium, formed from the hardened and darkened cuticle of the third instar larva. Because the puparium is an opaque structure that shows virtually no external changes, qualitative analyses of the internal tissues can be very informative for determining reliable age-specific morphological markers. Those analyses can be performed using either non-invasive but expensive and not widely accessible techniques, or traditional histological methods, which are invasive as they require the serial sectioning of the sample. Histological methods are often readily available for forensic researchers and practitioners; however, the histological study of blow fly intra-puparial stages has traditionally been hampered by the poor paraffin infiltration of tissues due to the abundance of fat bodies, resulting in usually fragmented sections and the subsequent loss of relevant information. We present here an effective method for the preparation of histological sections of blow fly intra-puparial stages, maximising the paraffin infiltration while enabling the production of clean and entire sections that allow for the use of reliable age-specific morphological markers, thus improving the accuracy of minPMI estimations when access to more costly techniques is not feasible.

Succession of medico-legal important flesh flies (Diptera: Sarcophagidae) in the temporal gradient of pig decomposition in the Brazilian Cerrado

Flies belonging to the Sarcophagidae family play a significant role in forensic investigations by aiding in the estimation of post-mortem interval through the assessment of the developmental time of their immature stages and/or the succession patterns of species on carcasses and cadavers. In this regard, this study aimed to investigate the ecological succession of these flies on pig carcasses within Cerrado of Brazil. The progression of fly succession was examined over a 10-day period using the Threshold Indicator Taxa Analysis (TITAN) approach. Six Z + species (Oxysarcodexia thornax (Walker), Peckia (Sarcodexia) lambens (Wiedemann), Peckia (Euboettcheria) collusor (Curran & Walley), Ravinia belforti (Prado & Fonseca), Tricharaea (Sarcophagula) canuta (Wulp) and Tricharaea (Sarcophagula) occidua (Fabricius)) were identified, with change points ranging from 2.5 to 3 days during the dry season and 2.5 to 5.5 days during the rainy season. Two Z−  species (Dexosarcophaga carvalhoi (Lopes) and Peckia (Sarcodexia) tridentata (Hall)) were present, with a change point of 6.5 days during the rainy season. This study provides a continuous analysis of the temporal succession of flies, enabling an investigation into species progression based on their change points and directions (Z + and Z− ), thereby offering methodological advancements to avoid the arbitrary categorization of inherently continuous data.

‘Not everything that can be counted counts’ in ethanol toxicological results: an antemortem and postmortem technical interpretation focusing on driving under the influence

Abstract Ethanol blood analysis is the most common request in forensic toxicology, and some studies point to positive results in approximately one-third of all unnatural deaths. However, distinguishing sober deaths from drunk deaths is not as simple as it may seem. This technical, clinical, and forensic interpretation is proposed to interpret the ethanol toxicological results, discussing several artifacts and pitfalls that must be considered, namely focusing on driving under the influence. This work is presented with a practical and objective approach, aiming to alleviate the complexities associated with clinical, physiological, pathophysiological, and toxicological aspects to enhance comprehension, practicality, and applicability of its content, especially to courts. Particularly the physical integrity of the body, the postmortem interval, putrefactive signs, anatomic place of blood collection, alternative samples such as vitreous humor and urine, the possibility of postmortem redistribution, the inclusion of preservatives in containers, and optimal temperature conditions of shipment are among some of the aspects to pay attention. Although the proposal of several biomarkers of postmortem microbial ethanol production, their translation to forensic routine delays is limited due to current uncertainties and analytical constraints that render their application. Specifically, in the interpretation of ethanol toxicological results, ‘not everything that can be counted counts and not everything that counts can be counted’ (attributed to Albert Einstein).

Estimation of Post-mortem Interval Based on Livor Mortis using a Colorimeter in Thai Populations

OBJECTIVE Livor mortis is a helpful and widely used method of estimating postmortem interval (PMI) in Thailand. This study aimed to investigate the value of a colorimeter as a tool for estimating the PMI. METHODS The color of livor mortis and control skin in 80 cadavers whose PMI was within 12 hours was measured by a colorimeter. The L* (brightness), a*b* (chroma and hue), and ∆E* values were compared to the control skin values. Statistical analysis was performed to determine the relationship between PMI and skin color before and after application of a specific pressure. RESULTS The results showed that colorimetric parameters were only weakly correlated with the PMI. An univariable analysis of ∆E* values was performed and showed good discriminatory power, with an area under the ROC curve of 0.82. The recommended cut-off value of ∆E* was 14 for the discrimination between early PMI (less than 6 hours) and late PMI (6-12 hours), in which the sensitivity and specificity were 72.5% and 80%. CONCLUSIONS The findings in this study reinforce the utility of colorimetric measurements in PMI estimation. With additional study and a larger sample size, the estimation of PMI could be established for general use in forensic practice.

CT ANALYSIS OF SKULL CONTENTS IN NATURALLY MUMMIFIED HUMAN CORPSES, A MULTICENTRIC STUDY.

This study aimed to assess skull contents, brain appearance, and density on postmortem computed tomography in naturally mummified corpses. For this purpose, a retrospective multicentric study, including mummified corpses from two French centers (Brest and Nantes) and from the New Mexico Decedent Image Database (USA), was performed by analyzing postmortem computed tomography (PMCT) focused on the head and neck of partially or fully mummified corpses discovered between 2011 and 2022. The PMCT analysis provided data on the CT appearance of brains, allowing them to be classified into four different categories (desiccation, liquefaction, dura mater only (DMO), and absence), and to measure densities (HU) of the brain remains. In addition, data on post-mortem intervals (PMI) from Nantes and Brest centers were collected and analyzed to test the link between brain densities and PMIs. 54 cases of naturally mummified corpses were included. The brains were classified as liquefied (56%), desiccated (17%), DMO (20%), and absent (7%) based on their CT appearance. Dehydrated brains were significantly (p<0.004) denser (median 102 HU, interquartile range (IQR) 41) than either liquefied brains (median 39.5 HU, IQR 9) or brains with DMO (median -25 HU, IQR 57). However, the density of brain remains was not significantly affected by where the bodies were found (p = 0,41). Analysis of PMI and brain densities was performed on 22 cases. The results showed that brain remains were significantly (p = 0.039) denser when they were found after a PMI of more than six months. Brain desiccation was the aspect with the highest densities on PMCT, and for which we were able to highlight great preservation of anatomical structures observable in living organisms.

Availability of individual proteins for quantitative analysis in postmortem brains preserved in two different brain banks.

Postmortem brain research is necessary for elucidating the pathology of schizophrenia; an increasing number of studies require a combination of suitable tissue samples preserved at multiple brain banks. In this study, we examined whether a comparative study of protein expression levels can be conducted using postmortem brain samples preserved in different facilities. We compared the demographic factors of postmortem brain samples preserved in two institutions and measured and compared the expression levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and glial fibrillary acidic protein (GFAP) in the prefrontal cortex and superior temporal gyrus. GAPDH is generally used as a loading control for western blotting, and GFAP is considered as an astrocyte marker in the brain. We found significant differences between the two institutions in postmortem interval, age at death, and preservation time. To reduce the effects of these differences on our measurements, the parameters were set as covariates in our analyses of covariance. Subsequently, no differences in GAPDH and GFAP expression were found between institutions. When studies are conducted using brain samples preserved in different brain banks, differences in demographic factors should be carefully considered and taken into account by statistical methods to minimize their impact as much as possible. Since there was no significant difference in the protein expression levels of GAPDH and GFAP in either region between the two institutions that preserved the postmortem brains, we concluded that it is possible to perform protein quantitative analysis assuming that there is no effect of difference between two institutions.

Research autopsy programmes in oncology: shared experience from 14 centres across the world.

While there is a great clinical need to understand the biology of metastatic cancer in order to treat it more effectively, research is hampered by limited sample availability. Research autopsy programmes can crucially advance the field through synchronous, extensive, and high-volume sample collection. However, it remains an underused strategy in translational research. Via an extensive questionnaire, we collected information on the study design, enrolment strategy, study conduct, sample and data management, and challenges and opportunities of research autopsy programmes in oncology worldwide. Fourteen programmes participated in this study. Eight programmes operated 24 h/7 days, resulting in a lower median postmortem interval (time between death and start of the autopsy, 4 h) compared with those operating during working hours (9 h). Most programmes (n = 10) succeeded in collecting all samples within a median of 12 h after death. A large number of tumour sites were sampled during each autopsy (median 15.5 per patient). The median number of samples collected per patient was 58, including different processing methods for tumour samples but also non-tumour tissues and liquid biopsies. Unique biological insights derived from these samples included metastatic progression, treatment resistance, disease heterogeneity, tumour dormancy, interactions with the tumour micro-environment, and tumour representation in liquid biopsies. Tumour patient-derived xenograft (PDX) or organoid (PDO) models were additionally established, allowing for drug discovery and treatment sensitivity assays. Apart from the opportunities and achievements, we also present the challenges related with postmortem sample collections and strategies to overcome them, based on the shared experience of these 14 programmes. Through this work, we hope to increase the transparency of postmortem tissue donation, to encourage and aid the creation of new programmes, and to foster collaborations on these unique sample collections. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Effects of antibiotics ceftriaxone and levofloxacin on the growth of Protophormia terraenovae (Diptera: Calliphoridae).

Protophormia terraenovae is a colonizer of decomposing bodies and is known to cause pre-mortem myiasis as the female flies lay eggs in uncleaned wounds. In this study the effects of different concentrations of antibiotics levofloxacin and ceftriaxone on maggot development, weight, length, and mortality were examined. The maggot length and weight were significantly increased by therapeutical doses of levofloxacin and ceftriaxone. The maggot development time was significantly decreased in every levofloxacin treatment compared to the control. The time to start pupation was significantly increased in the control compared to the antibiotic treatments. Levofloxacin significantly increased the survivability of the maggots. Every levofloxacin treatment significantly improved the rearing conditions for the maggots. Reaching the third instar was delayed by 24h in the control compared to the Levo 3.57 treatment. The Pupation in the control was delayed by an average of 48h compared to the Levo 3.57 treatment. The significantly reduced development time of the maggots in the antibiotic treatments might lead to an overestimation of the post-mortem interval and therefore an incorrect time of death determination. The improved rearing conditions may be an indication of the potential of a combined application of antibiotics and maggot therapy.

Abstract 2926: The effect of cancer research autopsy parameters on DNA and RNA sequencing quality

Abstract Background: Cancer research autopsy genomic studies offer insight into the metastatic cancer landscape but come with complexities that relate to the sampling and processing of post-mortem tissue. Clarifying the effect of autopsy variables on pre- and post-sequencing quality control (QC) is an unmet need that may inform tissue collection strategies. Methods: The effect of age, sex, post-mortem interval (PMI), and sample type (primary, metastatic, or normal) on pre-sequencing QC (nucleic acid concentration and integrity) was examined in 2678 samples (88.6% metastatic, 8.0% primary, 3.4% normal) from 83 patients with melanoma, lung, renal, or prostate cancer in the PEACE study. In the lung cohort, 160 surgical samples were also included through the TRACERx study, allowing surgery-autopsy tissue comparisons. Post-sequencing QC metrics were evaluated for lung samples that underwent DNA (n=522) or RNA (n=366) sequencing. Results: RNA concentration and RIN were greater in surgical samples than those collected at autopsy. Across cohorts, metastatic autopsy samples had greater nucleic acid concentrations than primary or normal autopsy samples, but not integrity. DNA and RNA concentration and integrity differed significantly between primary tumor types. When comparing samples of different metastatic sites from the whole cohort, concentration was lowest in bone (DNA) or the digestive tract (RNA), while integrity was greatest in the brain and lowest in the digestive tract (DIN, RIN). Although autopsy variables like age, sex and PMI correlated with pre-sequencing QC metrics in univariate analysis, they were not found to significantly correlate with these metrics in multivariate analysis, which identified that only primary cancer type and metastatic site were independent determinants of concentration and integrity. Similarly, for post-DNA (whole exome) sequencing QC, only the metastatic site was found to independently influence sequencing QC metrics like total number of sequences, average sequence length, and FastQC score. For RNA sequencing, only the metastatic site was found to influence sequencing QC metrics like total number of sequences, percentage of non-duplicated sequences, one hit-one genome percentage, and the alignment percentage on the human genome. Discussion: The lack of influence of PMI on QC in the largest QC-focused autopsy cancer study to date suggests that quality tissue can be obtained from non-rapid autopsy programs, which are more feasible and less resource-intensive than rapid programs. Citation Format: Petros Fessas, Sonya Hessey, Corentin Richard, Cristina Naceur-Lombardelli, Sophia Ward, David A. Moore, Karolina Nowakowska, Blanca Trujillo, Irene Lobon, Scott T. Shepherd, Fiona Byrne, Samra Turajlic, Gerhardt Attard, Charles Swanton, Mariam Jamal-Hanjani. The effect of cancer research autopsy parameters on DNA and RNA sequencing quality [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2926.

The micro CT evaluation of crown and root pulp volume versus dentin thickness in teeth in postmortem interval (PMI).

Determining the postmortem interval (PMI) is one of the main study subjects of forensic sciences. The main purpose of this prospective in vitro study that was the Micro-CT evaluation of teeth crown and root pulp volume versus dentin thickness in terms of PMI determination. The study involved 60 female Wistar rats, with weights ranging from 270 to 320g. These rats were grouped into six different post-mortem period categories. Following the animals' sacrifice, they were subjected to a natural putrefaction period, with a control group, in the grounds of a sheltered garden. Hemi-mandible samples were then extracted and placed in glass tubes for Micro-CT evaluations, following the progression of putrefaction processes. The pulp volume and dentin thickness were assessed using Micro-CT, and the gathered data underwent statistical analysis. Micro-CT was employed to analyze sixty right mandibular second molar teeth in the hemi-mandible. The crown pulp volume exhibited a reduction in group 6, with a value of 0.239 mm3 after a three-month period of natural putrefaction (p < 0.001). There is statistically differences among groups in case of pairwise comparison (p < 0.05). However, the root pulp volume and dentin thickness variables did not display any statistically significant changes. Despite certain limitations associated with this study, the Micro-CT findings concerning teeth pulp volume can serve as an objective parameter, especially for late postmortem investigations and the estimation of time of death.

Using surface-enhanced Raman spectroscopy to probe artificial dye degradation on hair buried in multiple soils for up to eight weeks

The discovery of clandestine burials poses unique challenges for forensic specialists, requiring diverse expertise to analyze remains in various states. Bones, teeth, and hair often endure the test of time, with hair particularly exposed to the external environment. While existing studies focus on the degradation of virgin hair influenced by soil pH and decomposition fluids, the interaction between artificial dyes on hair and soil remains underexplored. This paper introduces a novel approach to forensic hair analysis that is based on high-throughput, nondestructive, and non-invasive surface-enhanced Raman spectroscopy (SERS) and machine learning. Using this approach, we investigated the reliability of the detection and identification of artificial dyes on hair buried in three distinct soil types for up to eight weeks. Our results demonstrated that SERS enabled the correct prediction of 97.9% of spectra for five out of the eight dyes used within the 8 weeks of exposure. We also investigated the extent to which SERS and machine learning can be used to predict the number of weeks since burial, as this information may provide valuable insights into post-mortem intervals. We found that SERS enabled highly accurate exposure intervals to soils for specific dyes. The study underscores the high achievability of SERS in extrapolating colorant information from dyed hairs buried in diverse soils, with the suggestion that further model refinement could enhance its reliability in forensic applications.

Determination of Postmortem Interval in Mice.

Despite the major use of mice in biomedical research, little information is available with regard to identifying their postmortem changes and using that information to determine the postmortem interval (PMI), defined as the time after death. Both PMI and environmental conditions influence decomposition (autolysis and putrefaction) and other postmortem changes. Severe decomposition compromises lesion interpretation and disease detection and wastes limited pathology resources. The goal of this study was to assess postmortem changes in mice in room temperature cage conditions and under refrigeration at 4 °C to develop gross criteria for the potential value of further gross and histologic evaluation. We used 108 experimentally naïve C57BL/6 mice that were humanely euthanized and then allocated them into 2 experimental groups for evaluation of postmortem change: room temperature (20 to 22 °C) or refrigeration (4 °C). PMI assessments, including gross changes and histologic scoring, were performed at hours 0, 4, 8, and 12 and on days 1 to 14. Factors such as temperature, humidity, ammonia in the cage, and weight change were also documented. Our data indicates that carcasses held at room temperature decomposed faster than refrigerated carcasses. For most tissues, decomposition was evident by 12 h at room temperature as compared with 5 d under refrigeration. At room temperature, gross changes were present by day 2 as compared with day 7 under refrigeration. Mice at room temperature lost 0.78% of their baseline body weight per day as compared with 0.06% for refrigerated mice (95% CI for difference 0.67% to 0.76%, P < 0.0005). This study supports the consideration of temperature and PMI as important factors affecting the suitability of postmortem tissues for gross and histologic evaluation and indicates that storage of carcasses under refrigeration will significantly slow autolysis.

Age Determination of Chrysomya megacephala Pupae through Reflectance and Machine Learning Analysis.

Estimating the age of pupa during the development time of the blow fly Chrysomya megacephala (Diptera: Calliphoridae) is of forensic significance as it assists in determining the time of colonization (TOC), which could help to determine the postmortem interval (PMI). However, establishing an objective, accurate, and efficient method for pupa age inference is still a leading matter of concern among forensic entomologists. In this study, we utilized hyperspectral imaging (HSI) technology to analyze the reflectance changes of pupa development under different temperatures (15 °C, 20 °C, 25 °C, and 30 °C). The spectrograms showed a downtrend under all temperatures. We used PCA to reduce the dimensionality of the spectral data, and then machine learning models (RF, SVR-RBF, SVR-POLY, XGBR, and Lasso) were built. RF, SVR with RBF kernel, and XGBR could show promise in accurate developmental time estimation using accumulated degree days. Among these, the XGBR model consistently exhibited the most minor errors, ranging between 3.9156 and 7.3951 (MAE). This study has identified the value of further refinement of HSI in forensic applications involving entomological specimens, and identified the considerable potential of HSI in forensic practice.

Postmortem sympathomimetic iris excitability

Background:A reliable estimation of time since death can be important for the law enforcement authorities. The compound method encompassing supravital reactions such as the chemical excitability of the iris can be used to further narrow intervals estimated by temperature-based methods. Postmortem iris excitability was mostly assessed by parasympatholytic or parasympathomimetic substances. Little is known regarding sympathomimetic agents. The present study aims to describe the postmortem iris excitability using the sympathomimetic drug phenylephrine. Methods:Cadavers were included after body donors gave written informed consent during lifetime. Exclusion criteria were known eye disease, or a postmortem interval exceeding 26 hours. A pupillometer with a minimum measurement range of 0.5 mm was used to determine the horizontal pupil diameter before and 20 minutes after the application of phenylephrine. Increase in pupil diameter was labeled as positive reaction, unchanged pupil diameter was labeled as negative reaction, and decrease in pupil diameter was labeled as paradox reaction. Results:30 eyes from 16 cadavers (median age = 80.0; 9 males, 7 females) were examined. Initial pupil size was in median 3.5 mm (interquartile range [IQR]: 3.0–4.5 mm) and progressed to 4.0 mm (IQR: 3.5–5.0 mm) 20 minutes after drug instillation. The achieved pupil diameter difference comprised in median 0.5 mm (IQR: 0.0–1.0 mm). A positive reaction was observed in 21 cases. Negative reactions were observed in 5 cases and paradox reactions in 4 cases. Overall, there was a statistically significant difference in diameter between the initial and the reactive pupil (P = 0.0002). Conclusion:Although relatively rarely used, sympathomimetic drugs seem to be eligible for chemical postmortem iris excitability. Currently, assessment of postmortem iris excitability usually only involves parasympatholytic and parasympathomimetic agents. The findings of the present study give a hint that the application of a third agent with a sympathomimetic mechanism of action could provide additional information. Further studies assessing such a triple approach in the compound method in comparison with the current gold standard for estimation of time since death are mandatory to ensure reliable results.

Volabolomic Fingerprinting for Post-Mortem Interval Estimation: A Novel Physiological Approach.

Death is a multifaceted process wherein each individual cell and tissue has a metabolic homeostasis and a time of functional cessation defined by the dying process as well as by intrinsic and extrinsic factors. Decomposition is physiologically associated with the release of different types of volatile organic compounds (VOCs), and these form volaboloma mortis. The main purpose of this study was to record the volabolomic fingerprint produced by volatile molecules during the physiological decomposition process of human tissue and muscle cells. The volatile chemical signature has important implications for an open issue in forensics and pathology, namely the estimation of the postmortem interval (PMI), which decreases in accuracy with the passage of time. Volatile metabolites emitted from human tissues and muscle cells at 0, 24, 48, and 72 h were recorded in real time with an electronic nose sensor device. The key findings were the continuous sampling of VOCs emitted from tissues and cells. These showed a common behavior as time progressed; particularly, after 48 h the distributions became dispersed, and after 72 h they became more variable. Volabolomic fingerprinting associated with time progression relevant to the study of PMIs was reconstructed. Additionally, there may be broader applications, such as in dog training procedures for detecting human remains, and perhaps even for studying scavenger and insect attractants.

Morphological changes of larvae and pupae of Lucilia sericata (Diptera: Calliphoridae) reared at two temperatures and on three food types.

Determining the minimum postmortem interval (minPMI) from an entomological perspective relies mainly on development data recorded for various species of flies collected from a crime scene or suspicious death. This study focused on the larval and pupal development of Lucilia sericata (Meigen), with an emphasis on the changes of the external morphology of the puparium and its pupal content throughout the duration of metamorphosis. Colonies of L. sericata were reared on 3 types of swine tissue (skeletal muscle, liver tissue, and heart tissue) at 2 different temperature regimes; 24 ± 1 °C and 30 ± 1 °C. The overall developmental time, larval width and length, and inner and outer pupal morphology changes were observed and recorded. The results show that: (i) temperature significantly influenced overall development time, as well as changes in larval width and length, but this effect was not dependent on tissue type; (ii) larval development duration was longest on heart tissue, and shortest on skeletal muscle for both temperatures; and (iii) pupation was longest for larvae reared on skeletal muscle at 24 ± 1 °C, and on liver tissue at 30 ± 1 °C, while those larvae reared on liver tissue at 24 ± 1 °C and heart tissue at 30 ± 1 °C had the shortest pupation period. A seven-character checklist plus 4 landmark stages were developed comprising the external morphology of the puparium and pupal content changes of L. sericata. In conclusion, the study provides larval and pupal development timetables, as well as checklists and photo guides for pupal character development that may be useful for future postmortem determinations.

Evaluation of porcine decomposition and total body score (TBS) in a central European temperate forest.

The total body score (TBS) is a visual scoring method to scale the succession of decomposition stages. It compares decomposition between cadavers, to connect it with external taphonomic factors and estimate the post-mortem interval. To study decomposition in various climatic environments, pigs are often used as human proxies. Currently, there is one TBS system by Keough etal. (J Forensic Sci. 2017;62:986) for surface-deposited domestic pigs, coming from South Africa. Our study aims to evaluate this method and analyze porcine decomposition in Central Europe to inform forensic research and casework. We conducted an experiment studying six 50 kg pig carcasses in a temperate Swiss forest. Three observers documented decomposition patterns and rated the decomposition stages from photographs based on the porcine TBS model by Keough etal. (J Forensic Sci. 2017;62:986). We documented discrepancies between the carcass decomposition of our specimens and those in the South African study, especially related to the high insect activity in our experiment. Furthermore, we noted factors complicating TBS scoring, including rainfall and scavengers. The agreement between TBS observers from photographs was in the highest agreement category apart from one "substantial agreement" category. Our study is the first in Europe to systematically test the Keough etal. (J Forensic Sci. 2017;62:986) method. The results evidence that regional adaptations are required to be applicable for other environments. We present a modified approach based on experimental observations in a Swiss temperate forest. The identification of regional decomposition patterns and drivers will inform future taphonomy research as well as forensic casework in comparable contexts in Central Europe.

Ante- and Post-Mortem Fracture Identification Protocol Based on Low- and High-Level Fusion Using Fourier Transform Infrared Spectroscopy and Raman Spectroscopy Association.

In this study, the application of low-level fusion (LLF) and high-level fusion (HLF) strategies using a combination of Fourier transform infrared spectroscopy (FT-IR) and Raman spectroscopy in the identification of antemortem and postmortem fracture at different postmortem intervals (PMIs) was investigated. On a technical level, the same hard tissue sample can be detected using a mix of FT-IR and Raman techniques. At the method level, two cutting-edge chemometrics approaches (LLF and HLF) combining FT-IR and Raman spectroscopic data are explored. The models were ranked in accordance with their parametric quality as follows: HLF and LLF + HLF models > LLF single model > Raman single model > FT-IR single model. The LLF model performed marginally better than the Raman model, however, when compared to other models, the HLF model performed considerably better. The HLF model achieved the best performance, with both cross-validation accuracy and test data set accuracy of 0.88. The importance of the feature wavelengths in the model construction process was subsequently evaluated by intersection fusion, and it was found that the absorbance bands of amide I, PO43- ν1 ν3, and CH2 in FT-IR and phenylalanine, CO32- ν1- PO43- ν3, and amide III in Raman have outstanding contributions to the construction of antemortem and postmortem fractures identification models. Overall, the combination of FT-IR and Raman with the HLF strategy is a novel and promising approach for developing antemortem and postmortem fracture identification models at different PMIs.

Improving accuracy of age estimates for insect evidence-calibration of physiological age at emergence (k) using insect size but without "k versus size" model.

Postmortem interval may be estimated based on the age of insect evidence collected on a death scene. Reference data that are used in such estimation frequently comprise thermal summation constant (i.e. k), which is equal to the insect age upon completion of immature development expressed in accumulated degree-days or degree-hours (ADD or ADH). Essentially, k is a central point of an insect group and it may poorly represent insect evidence that is near the limits of variation for the group. Accordingly, it was postulated to calibrate k for particular insect evidence and insect size and sex were found to be useful for this purpose in some of the species. However, the calibration is only possible by using the model that correlates k with insect size. Since very few such models were published, this lack of data limits the calibration of k in forensic casework. In this article, we develop a formula that is useful for the calibration of k without the use of "k versus size" model (and related datasets). The formula uses k from the general thermal summation model for a species (with its standard error), size range for the species (retrieved from entomology literature), and size measurements for particular insect evidence. The calibration of k with the formula was validated using the Creophilus maxillosus (Coleoptera: Staphylinidae) and Necrodes littoralis (Coleoptera: Silphidae) datasets. It was particularly useful while analyzing unusually small and large insects, in case of which the formula reduced the inaccuracy of k from the general model on average by ~25 ADD in C. maxillosus and ~40 ADD in N. littoralis. We discuss the limitations and prospects of the calibration protocol that employs the formula.