Andrea Bonicelli

Considering the growing importance of microbiome analyses in forensics for identifying individuals, this study explores the transfer of the skin microbiome onto clothing, its persistence on fabrics over time, and its transferability from the environment and between different garments. Furthermore, this project compares three specific QIAGEN microbiome extraction kits to test their extraction efficiency on fabric samples. Additionally, this study aims to check if these extracts contain human DNA, providing a chance to obtain more information from the same evidence for personal identification. The results obtained show: (1) variations in the skin microbiome between the volunteers, potentially due to their different sex; (2) differences in microbial composition between worn and unworn clothing; (3) the influence of the environment on the microbial signature of unworn clothing; (4) the potential use of certain phyla as biomarkers to differentiate between worn and unworn garments, even over extended periods; (5) a tendency towards extraction biases in the QIAampMP^® DNA microbiome kit among the three tested ones; and (6) none of the extraction kits allow for the typing of human genetic profiles suitable for comparison. In conclusion, our study offers supplementary insights into the potential utility of time-transferred microbiome analysis on garments for forensic applications.

Assessing the timing of skeletal trauma significantly impacts the reconstruction of events surrounding death and deposition in forensic cases. However, there are no absolute time frames in which the characteristics of wet bone (peri-mortem) fractures transition to dry (post-mortem) fractures. The aim of this study was to attempt to identify a point within the post-mortem interval in which the characteristics of bone change from wet to dry bone properties. A total of 32 deer ribs were placed in a laboratory burial environment and a set of three were fractured with blunt force trauma every week during a ten-week period. All samples and the inflicted trauma effects were documented and analysed by macroscopic observation, scanning electron microscope (SEM) analysis, thermal analysis, biomechanical analysis, and attenuated total reflectance–Fourier transform infrared spectroscopy (ATR-FTIR). No significant difference was found in the macroscopic, microscopic, thermal, and biomechanical analyses of the trauma inflicted over the 10-week period. A significant difference was only found in the carbonate-to-phosphate ratio in analytical chemistry. The results suggest that interpreting wet bone characteristics in forensic anthropology as having been inflicted during the peri-mortem period (around the time of death) should also consider that these, in fact, could be inflicted well after death (post-mortem) as wet bone properties as this study has shown persist at least 10 weeks after death in a burial environment.

The combined use of multiple omics allows to study complex interrelated biological processes in their entirety. We applied a combination of metabolomics, lipidomics and proteomics to human bones to investigate their combined potential to estimate time elapsed since death (i.e., the postmortem interval [PMI]). This ‘ForensOMICS’ approach has the potential to improve accuracy and precision of PMI estimation of skeletonized human remains, thereby helping forensic investigators to establish the timeline of events surrounding death. Anterior midshaft tibial bone was collected from four female body donors before their placement at the Forensic Anthropology Research Facility owned by the Forensic Anthropological Center at Texas State (FACTS). Bone samples were again collected at selected PMIs (219-790-834-872days). Liquid chromatography mass spectrometry (LC-MS) was used to obtain untargeted metabolomic, lipidomic, and proteomic profiles from the pre- and post-placement bone samples. The three omics blocks were investigated independently by univariate and multivariate analyses, followed by Data Integration Analysis for Biomarker discovery using Latent variable approaches for Omics studies (DIABLO), to identify the reduced number of markers describing postmortem changes and discriminating the individuals based on their PMI. The resulting model showed that pre-placement metabolome, lipidome and proteome profiles were clearly distinguishable from post-placement ones. Metabolites in the pre-placement samples suggested an extinction of the energetic metabolism and a switch towards another source of fuelling (e.g., structural proteins). We were able to identify certain biomolecules with an excellent potential for PMI estimation, predominantly the biomolecules from the metabolomics block. Our findings suggest that, by targeting a combination of compounds with different postmortem stability, in the future we could be able to estimate both short PMIs, by using metabolites and lipids, and longer PMIs, by using proteins.

Histomorphometry constitutes a valuable tool for age estimation. Histological interpopulation variability has been shown to affect the accuracy of age estimation techniques and therefore validation studies are required to test the accuracy of the pre-existing methodologies. The present research constitutes a validation study of widely known histological methods on the sixth rib and the femoral midshaft of a 19th century British population originating from Blackburn, England. An evaluation of the histomorphometric features of eleven ribs and five femora was performed and used to test the accuracy of selected methods. Results indicated that osteon area and circularity were the only histomorphometric variables that presented significant interpopulation variability. Cho et al.’s method for the ribs and the average value produced using Kerley and Ubelaker’s method for intact osteon and percentage of lamellar bone equations for femur were considered the only reliable markers for estimating the age on the Blackburn sample. In the case of old individuals, Goliath et al.’s method provided more satisfactory results. Overall, the present study provides evidence on the applicability of the aging histomorphometric methods on a British sample and highlights the limitations of applying histomorphometric methods developed on different reference populations than the one under investigation.

The study of post-mortem changes is a crucial component of forensic investigation. Human forensic taphonomic facilities (HFTFs) are the only institutions allowing the design and execution of controlled human decomposition experiments. When bodies are skeletonized, bones are normally stored in skeletal collections and used for anthropological studies. However, HFTFs apply chemical and/or thermal treatments to the remains prior bone long-term storage. These treatments are believed to alter heavily the original biochemical and molecular signature of bone material. The present study aims to evaluate the effect of these procedures on the bone metabolome and lipidome by using an animal bone model. Three intact bovine tibiae were processed using three protocols routinely applied at HFTFs, and their three counterparts were used as non-treated controls. Bone powder samples were subjected to biphasic extraction and both metabolites and lipids were analysed via liquid chromatography tandem mass-spectrometry. Results showed severe reductions in the abundances of both metabolites and lipids, and the presence of contamination introduced by cleaning agents. Despite the preliminary nature of the study, we demonstrated that the biochemical profile of bone is heavily affected by the maceration procedures. Ideally, these treatments should be avoided, or replaced by minimally invasive procedures agreed across HFTFs.

Bone-mineral density (BMD) is a measure of the inorganic mineral content in bone, and is one of the more informative assessments of bone quality in both clinical studies and forensic investigations. Several factors, such as age, sex, disease, genetics, and lifestyle, affect BMD measurements, and normative standards must be applied for specific groups and individuals. One of the most common disorders associated with low BMD is osteoporosis and increased fracture risk, due to a decrease in bone strength and an increase in bone fragility. Medical conditions like diabetes or hyperthyroidism and other parameters like peak bone mass and postmenopausal estrogen deficiency also impact BMD. Single- and dual-energy photon absorptiometry, quantitative computet tomography, and magnetic resonance imaging are some of the technological modalities for BMD quantification, and each presents distinct advantages and limitations, depending on the purpose of the analysis, the specific characteristics of the individual, the bone site under examination, and the equipment and trained personnel available. Recently, BMD values were applied to forensic medicine in a variety of scenarios ranging from age and sex estimation to the assessment of malnutrition and the use of finite-element modelling. Despite technical and methodological inconsistencies reported in the literature on BMD readings, there is scope for expanding the use of this variable in forensic settings.