Skin surface debris as an archive of environmental traces: an investigation through the naked eye, episcopic microscope, ED-XRF, and SEM-EDX.

Traces from bodies can be of various nature, for example of biological or inorganic origin. Some of these historically have received more consideration than others in forensic practice. Samplings of gunshot residues or biological fluid traces are commonly standardized, whereas macroscopically invisible environmental traces are usually ignored. This paper simulated the interaction between a cadaver and a crime scene by placing skin samples on the ground of five different workplaces and inside the trunk of a car. Traces on samples were then investigated through different approaches: the naked eye, episcopic microscope, Scanning Electron Microscopy (SEM) with Energy-dispersive X-ray spectroscopy (EDX) and Energy Dispersive X-Ray Fluorescence (ED-XRF). The purpose is to provide the forensic scientist with the awareness of the value of debris on skin and then to highlight implications for forensic investigations. Results demonstrated that even naked eye observation can reveal useful trace materials, for defining the possible surrounding environment. As a next step, the episcopic microscope can increase the number of visible particulates and their analysis. In parallel, the ED-XRF spectroscopy can be useful to add a first chemical composition to the morphological data. Finally, the SEM-EDX analysis on small samples can provide the greatest morphological detail and the most complete chemical analysis, although limited, like the previous technique, to inorganic matrices. The analysis of debris on the skin, even with the difficulties due to the presence of contaminants, can provide information on the environments involved in criminal events that can add to the investigation framework.

Water-related atmospheric agents and solubility: two parameters of validation in toxicological screening on clothing worn by skeletal remains.

In forensic toxicology, when conventional matrices are no longer available, alternative matrices can be used to assess toxicological investigations. Clothes worn by skeletal remains may be a good unconventional matrix for toxicological analyses considering that they have absorbed decomposition fluids and blood from a body. We hypothesized a scenario in which a skeleton, wearing clothes, was discovered in an open environment. From this starting point, an experimental study was developed on different textiles (cotton, wool, and polyester) to evaluate whether water-related atmospheric agents and molecule solubility can largely influence the detection of molecules of toxicological interest on this specific matrix, together with the characteristics of different garments chosen. The experimental study was performed on blood spots, previously spiked with 6-monoacetylmorphine and morphine, accurately placed on different textiles and washed with different quantities of deionized water adjusted at pH 5.6 with formic acid to simulate different rainfall conditions. Toxicological analyses were performed via Solid-Phase Extraction and High-Performance Liquid Chromatography-Tandem Mass Spectrometry analyses (Thermo Scientific™ TSQ Fortis™ II Triple-Quadrupole Mass Spectrometer). From the experimental study morphine could not be detected on 100% cotton and 100% wool fabric after the passing of 500 mL of deionized water and in 100% synthetic polyester textile after washing with 250 mL of deionized water. In conclusion, when toxicological analyses are carried out on unconventional matrices as textiles worn by corpses exposed to different environmental conditions, it is of great importance, in using such substrates as evidence for the presence of molecules of toxicological interest, to evaluate chemical-physical characteristics of each analyte under investigation in order to correctly interpret the toxicological data obtained.

The rights of migrants to the identification of their dead: an attempt at an identification strategy from Italy.

Europe is turning a blind eye on a humanitarian disaster unfolding at its doorsteps, with thousands of migrants dying unidentified in Mediterranean waters. Since 2014, Italy has been struggling in an almost indifferent international scenario to identify its dead migrants. Despite the lack of sufficient resources, of the difficulties in collecting post mortem data from the disseminated bodies, and of the problems of contacting and collecting ante mortem information from relatives, it has been proven, with a series of pilot studies, that not only can these bodies be identified but that relatives are also looking for their loved ones and need death certificates. This article focuses on the administrative limbo and lack of regulations obliging single states to engage in appropriate procedures to maximise identification.

Traces under nails in clinical forensic medicine: not just DNA.

When dealing with complex crimes such as rape and assault, every trace takes on an essential role. The hands are often the only means of defence and offence for the victim as well as a frequent area of contact with the environment; fingernails of a victim are a well-known possible source of DNA of the aggressor; nevertheless, they are more rarely treated as an area of interest for non-genetic material, particularly on living victims. The hyponychium, because of its physiological protective function, lends itself ideally to retaining different kinds of traces representative of an environment or various products and substrates that could shed light on the environment and objects involved in the event. We therefore tested how far this capability of the hyponychium could go by simulating the dynamics of contamination of the nail through scratching on different substrates (brick and mortar, painted wood, ivy leaves, cotton and woollen fabric, soil) and persistence of any contaminant at different time intervals. We have thus shown how these traces may remain in the living for up to 24 h after the event using inexpensive and non-destructive techniques such as the episcopic and optical microscope.

Common and much less common scenarios in which botany is crucial for forensic pathologist and anthropologists: a series of eight case studies.

It is commonly accepted that crime scene recovery and recording are key moments of any judicial inspection in which investigators must decide on the correct strategies to put into place. Complex outdoor scenarios, presenting partially or entirely skeletonised remains, can benefit more than others by the intervention of environmental specialists (forensic anthropologists, archaeologists, entomologists and botanists). These experts are capable of singling out, correctly recording and recovering environmental evidence that can lead to a more comprehensive reconstruction of a given criminal episode. If human remains are discovered in an outdoor scenario, the on-site presence of a botanist will guarantee a correct approach to the identification, recording and recovery of any botanical evidence. If an on-site botanist is not available, the operators must be capable of both the botanical evaluation of a scene and the implementation of correct botanical sampling protocols.The following collection of unusual case histories that aim at underlining the efficacy of forensic botany will examine the determination of post mortem or the post depositional interval, evidence for a victim's post mortem transfer, evidence for the identification of a primary crime scene and evidence for the identification of a victim's dismemberment site. In another two cases, one, we will illustrate the important role that forensic botany played in the discrimination between botanical material used to voluntarily conceal a victim and vegetation that had grown naturally above a disposal site, whereas the other will highlight the protocols implemented for the identification of a murder weapon.

Histological determination of the human origin from dry bone: a cautionary note for subadults.

Anthropologists are frequently required to confirm or exclude the human origin of skeletal remains; DNA and protein radioimmunoassays are useful in confirming the human origin of bone fragments but are not always successful. Histology may be the solution, but the young subadult structure could create misinterpretation. Histological tests were conducted on femur and skull of 31 human subjects. Each sample was observed focusing on presence or absence of fibrous bone, lamellar bone, radial lamellar bone, plexiform bone, reticular pattern, osteon banding, Haversian bone, primary osteons, secondary osteon and osteon fragments. Samples were divided into five age classes; 1 (<1 year), 2 (1-5 years), 3 (6-10 years), 4 (11-15 years) and 5 (16-20 years). Regarding femurs, class 1 presented the following: 87.5% fibrous bone, 37.5% plexiform bone, 12.5% reticular pattern and 12.5% lamellar bone radially oriented. Class 2 showed 37.5% of fibrous bone, 12.5% of reticular pattern and 37.5% of osteon banding. In the higher age classes, the classical human structures, lamellar bone and osteons were frequently visible, except for one case of reticular pattern, generally considered a distinctive non-human structure. The situation appeared different for the skull, where there was a lack of similar information, both in human and non-human. An analysis of the percentage of lamellar bone and osteons was conducted on femur and skull fragments. A trend of increase of primary osteon number and a decrease of the lamellar bone area has been detected in the femur. The present study has therefore shed some light on further pitfalls in species determination of subadult bone.

Pathological alterations and COHb evaluations as tools for investigating fire-related deaths in veterinary forensic pathology.

Fire-related deaths are usually a consequence of carbon monoxide (CO) poisoning or shock from thermal injuries. In humans, high levels of carboxyhemoglobin (COHb) concentrations in the blood can support a diagnosis of CO poisoning. In veterinary medicine, few studies investigated the pathological changes and blood COHb% in fire victims, and no data are available on post-mortem changes in blood gas composition due to fire. This study aims to investigate the pathological changes and COHb levels in both animal victims of fire and cadavers experimentally exposed to fire. For this purpose, dogs were selected and subdivided into three groups. Group A comprised 9 adult dogs, and Group B comprised 7 puppies that died under fire-related conditions. Group C was represented by 4 dog cadavers experimentally exposed to heat and smoke. A complete macroscopic, histological, and COHb evaluation were performed on each animal. Animals in Groups A and B showed cherry-red discoloration, thermal-injuries and soot deposits along the respiratory tract. Animals in Group C showed thermal injuries and soot deposits limited to the upper respiratory tract. The mean COHb% values in cadavers in Group C were lower than those observed in the other groups but higher compared to the values detected before the heat and smoke treatment. These findings suggest that both pathological changes and COHb analysis are valid tools for investigating fire-related deaths in dogs. However, the increase of COHb levels in cadavers exposed post-mortem to heat and smoke highlights how the COHb analysis should always be evaluated together with macroscopical and microscopical findings to avoid significant misjudgments in investigating fire-related fatalities in veterinary forensic practice.

Evaluation of Muscle Proteins for Estimating the Post-Mortem Interval in Veterinary Forensic Pathology.

Postmortem cadaveric changes are commonly used to estimate the postmortem interval (PMI) in humans and animals. However, these modifications have been poorly investigated in animals of interest to veterinary forensic pathology. The aim of this study was to investigate the potential use of muscle proteins (desmin and dystrophin) as biomarkers for estimating the PMI in dogs. For this study, 10 dead adult dogs were evaluated for 4 days in a temperature-controlled room at 19 ± 1 °C. For each animal, at 3, 24, 48, 72, and 96 h after death, a 1 × 1 × 1 cm cube of muscle tissue was removed from the vastus lateralis and triceps brachii. Protein expression levels were analyzed by immunohistochemical examination and immunoblot analysis. The obtained results showed rapid dystrophin degradation, with complete disappearance at 72 h after death. In contrast, desmin-positive fibers and desmin protein bands detected by immunoblot were observed on all 4 days of observation. Our findings suggest the potential use of muscle proteins as biomarkers for estimating the PMI in dogs.

Skeletal Markers of Physiological Stress as Indicators of Structural Violence: A Comparative Study between the Deceased Migrants of the Mediterranean Sea and the CAL Milano Cemetery Skeletal Collection.

Structural violence is an indirect form of violence that can lead to physiological consequences. Interestingly, these physiological disruptions may affect the skeletons and can therefore provide relevant information on violence and way of life in the analysis of skeletal remains. The aim of the present study was to test the hypothesis that migrants who died in the Mediterranean Sea would present physiological cranial stress markers such as cribra orbitalia (CO), porotic hyperostosis (PH), and linear enamel hypoplasia (LEH) more frequently and more severely than Italians of the 20th century. With this intent, a total of 164 crania were examined: 139 from deceased migrants recovered from a shipwreck in the Mediterranean Sea in 2015, aged between 16 and 35 years old, and 25 of the same age from the CAL Milano Cemetery Skeletal Collection. Both presence and severity of CO, PH, and LEH were evaluated. The data obtained were analyzed using Wilcoxon signed-rank and independence Chi-squared tests to compare the results between the two samples and to test whether there was an association between the sample of migrants and the occurrence of lesions. As a result, CO and PH appeared more frequently and more severely in the migrant sample. In addition, migrants were significantly associated with CO, PH, and LEH (p-values < 0.05). Although this does not imply in any way that CO, PH, and LEH are specific to migration, they should be regarded as indicators of structural violence.

Comparing Genetic and Physical Anthropological Analyses for the Biological Profile of Unidentified and Identified Bodies in Milan.

When studying unknown human remains, the estimation of skeletal sex and ancestry is paramount to create the victim's biological profile and attempt identification. In this paper, a multidisciplinary approach to infer the sex and biogeographical ancestry of different skeletons, using physical methods and routine forensic markers, is explored. Forensic investigators, thus, encounter two main issues: (1) the use of markers such as STRs that are not the best choice in terms of inferring biogeographical ancestry but are routine forensic markers to identify a person, and (2) the concordance of the physical and molecular results. In addition, a comparison of physical/molecular and then antemortem data (of a subset of individuals that are identified during our research) was evaluated. Antemortem data was particularly beneficial to evaluate the accuracy rates of the biological profiles produced by anthropologists and classification rates obtained by molecular experts using autosomal genetic profiles and multivariate statistical approaches. Our results highlight that physical and molecular analyses are in perfect agreement for sex estimation, but some discrepancies in ancestry estimation were observed in 5 out of 24 cases.

Rediscovering the value of images in supporting personal identification of missing migrants.

The crisis concerning the identification of dead migrants is forcing forensic scientists to deal with the unavailability of classical antemortem (AM) data such as radiological information and DNA. However many times there is abundant AM photographic data of which the availability and quality is underestimated. Independently of when facial image comparison can be used as a valid identification method, description of the unexpected quality and quantity of images in the scenario of unidentified migrant decedents is given in this observational study. It focuses on the overall characteristics of the AM images available for a group of 74 missing migrants related to the disaster of October 3rd, 2013. 394 images were collected: an average of 6 for each missing person. 18% are original pictures, 47.5% scans of original images, 9.9% scans of printed images while 9.4% downloaded from social networks such as Facebook. 42.7% of images resulted in focus. Among all the images, about the 10% has been used in the personal identification process to ascertain the identity of 22 persons, because of the visibility of identifying details (facial, dental and body features). The importance of such a finding should not be diminished by its simplicity. Today, those involved in collecting AM information from relatives of dead migrants are underestimating the possible presence of images, even in sub-Saharan populations, where, contrary to belief and data, technology, though simple, is available and images may turn out to be abundant and useful (and at times the only resort) for identification.

Contribution of plant anatomy to forensic investigation: Tree bark morphology.

Plant science has been more and more utilized in forensic investigation, although its full potential is still to be reached. Plant macroremains are a powerful tool to link a body or other evidence back to a primary crime scene as they can provide detailed information about its previous ecological and geographic location. However, plant macroremains are often poorly preserved and difficult to identify, as diagnostic elements are seldom present within the assemblage occurring on the scene. Plant fragments most likely to be found are those exposed to the environment and resistant to degradation. The bark of woody plants meets these requirements but the possibility of its identification at species level from small fragments is not known. Starting from a real homicide case, where bark splinters were found on the victim, we aimed to assess the forensic potential of bark identification from small fragments like those likely to occur on a crime scene. Two identification keys were prepared for 16 common lowland tree species from Northern Italy; one key used all the available anatomical traits, the second only those from the outer bark. The second key was not able to discriminate some couples of species unambiguously, but could identify the bark fragments of the homicide as Robinia pseudoacacia, as confirmed from direct comparison with a reference sample. Bark fragments deserve to be included into the macroremains to be analyzed during an investigation, but small samples could easily lack diagnostic traits, and the building of a reference collection should be encouraged.

Look before washing and cleaning: A caveat to pathologists and anthropologists.

Partially or totally skeletonized bodies are undoubtedly the most challenging scenario to deal with for forensic pathologists and anthropologists. Indeed, in such cases, being able to figure out the cause and manner of death is often tricky. Human remains require to be washed and cleaned before a thorough assessment of any signs of trauma. However, bones and any fragment of more or less putrefied soft tissues may be home to crucial traces for investigative purposes. They are often located in the context of apparently meaningless dirt and, sometimes they are even invisible to the naked eye. Therefore, their careless cleaning inevitably leads to an unintentional loss of such traces with a negative impact on subsequent investigations. For these reasons before proceeding with cleaning, exhaustive examination and sampling must be carried out. In particular fragments of soft tissues, even if putrefied, are absolutely not to be considered as a hindrance for forensic purposes, since they could still provide valuable information after histological examination. Finally, forward-thinking professionals should think about the possible presence of exogenous micro-traces of forensic concern and collect specimens to be analyzed through in-depth analyses, such as Scanning Electron Microscopy with Energy Dispersive X-ray Spectrometry (SEM/EDX). The present series of cases demonstrates that crucial forensic information can be obtained through the analysis of apparently meaningless residues and even of micro-traces not visible to the naked eye and mixed with trivial dirt.

Effects of Cremation on Fetal Bones.

The charring process is a weak point of anthropological analysis as it changes bone morphology and reduces information obtainable, specially in fetuses. This experiment aims at verifying the conservation of fetal bones after cremation. A total of 3138 fetuses of unknown sex and age were used, deriving from legal and therapeutic abortions from different hospitals of Milan. Cremations took place in modern crematoria. Nine cremation events were analyzed, each ranging from 57 to 915 simultaneously cremated fetuses. During the cremations, 4356 skeletal remains were recovered, 3756 of which (86.2%) were morphologically distinguishable. All types of fetal skeletal elements were found, with the exception of some cranial bones. Only 3.4% of individuals could be detected after the cremation process, because of the prevalence of abortions under 12 lunar weeks. All fire alterations were observed and the results were statistically analyzed. This pilot study confirmed the possibility of preservation of fetal skeletal elements after cremation.