Could synovial fluid be a useful substrate for the forensic analysis of diatoms?

To date, synovial fluid has not been the subject of targeted analysis as a possible substrate to search for the presence of diatoms in the forensic context of drowning. However, its unique characteristics of production and isolation from the external environment could make it suitable for this purpose, similar to what has already been demonstrated in the literature for vitreous humor. By considering this, synovial fluid was analyzed in a specific case that came to our attention, where the coexisting signs of polytrauma and drowning were documented during autopsy, demonstrating a period of vitality during immersion. After a thin smear of the supernatant was obtained from the centrifugation of the synovial fluid sample, diatoms were successfully detected, consistent with those found in other organs and the water of the canal. The detection of diatoms in the synovial fluid was an objective finding, but its generalizability is limited because this was a pilot application. However, in cases where death by drowning is suspected and the body has multiple areas breached by trauma, the technique of analyzing diatoms in the synovial fluid could have great potential. Therefore, it is appropriate to further explore this technique in order to obtain more forensic evidence in such a setting.

Colorimetric Barium Detection of Gunshot Residues on Cadaveric Human Skin: A Pilot Application for Forensic Purposes.

Introduction: In this study we microscopically investigated, for the first time ever, the colorimetric detectability of barium of gunshot residues (GSR) on cadaveric human skin with gunshot wounds. Methods: For this purpose we used two different colorimetric techniques known in the literature, namely 0.2% sodium rhodizonate (Na-R-Ba 0.2%) and sodium rhodizonate in alcoholic environment (Na-R-Ba OH 0.2%). At the same time, we have also coupled it with scanning electron microscopy with energy dispersive X-ray (SEM/EDX) analysis and the colorimetric study for the detection of lead of GSR. These techniques were applied to 16 victims who died from gunshot injuries, as well as to a control group. Results: SEM/EDX demonstrated the presence of lead in all cases and barium in 11 of the 16 cases. The subsequent colorimetric technique with Na-R-Ba 0.2% did not show the barium of GSR in any case, unlike the Na-R-Ba OH 0.2% technique. This latter, in fact, has demonstrated the presence of this metal in 2 cases (18%). No microscopic case of false positive was recorded. Conclusion: The evidence obtained with Na-R-Ba OH 0.2% makes this method, applied here for the first time ever, worthy of further study. Meanwhile, although this technique can certainly be applied, it cannot be separated from the contextual colorimetric investigation for lead and the use of more sophisticated techniques.

Forensic pilot application of rehydrating solutions on human cadaveric skin: what are the effects on hemorrhagic infiltrates?

The microscopic evaluation of hemorrhagic infiltrates is crucial in forensic diagnostics, but it proves challenging in corificated and mummified cadavers. In these cases, pre-treatment with rehydrating solutions is recommended, although their effects on the hemorrhagic infiltrate are not well understood. In this pilot study, we microscopically investigated the effect of two different rehydrating solutions-Sandison's solution and fabric softener-on well-preserved human cadaveric skin samples taken from areas affected by an ecchymotic lesion, comparing them with direct fixation in formalin. Specifically, we examined the topographic distribution of the hemorrhagic infiltrate in each layer of the skin by assigning a semi-quantitative score, conducted mutual comparisons, and performed statistical analysis. Histologically, compared to direct fixation in formalin, a slight and statistically non-significant reduction in the hemorrhagic infiltrate was observed in samples pre-treated with fabric softener. On the other hand, a more pronounced and statistically significant decrease in scores was observed in samples pre-treated with Sandison's solution. This effect is likely due to the fact that Sandison's solution, due to its components, exerts an osmotic effect, partially inducing osmotic lysis of red blood cells. Overall, extensive areas of hemorrhagic infiltrates were preserved, although to a lesser extent, while smaller foci were markedly reduced, sometimes even disappearing. The findings suggest that Sandison's solution has a detrimental effect on cutaneous hemorrhagic infiltrates, emphasizing the importance of being cautious and conducting dual sampling, using both formalin and a rehydrating solution, for forensic examination of mummified or corificated skin samples.

Elastic fiber degradation as a possible indicator for PMI estimation on mummified and corified skin: a pilot study.

Mummified and corified bodies are particularly complex scenarios to investigate, starting from identifying the post-mortem interval (PMI), even more so in indoor environments. In these bodies, the skin has the peculiar feature to resist for a long time. Among its components, there are elastic fibers, which are characterized by intrinsic resistance to post-mortem degenerative phenomena. Starting from these considerations, we investigated microscopically the persistence, detectability, and changes of elastic fibers in the skin of mummified and corified bodies with different known PMI. The aim was to evaluate whether they could provide an additional tool to aid in PMI estimation in these cases. Therefore, we collected skin samples from mummified or corified bodies found in a domestic environment with different known PMI, as well as from corified bodies that had been exhumed after 11 years of burial. Histochemical staining specific for elastic fibers, namely, Weigert's resorcin fuchsin, showed their prolonged persistence and a progressive and different degradation between mummified and corified skin as a function of PMI. Moreover, on the whole, we observed greater preservation of elastic fibers in mummified skin than in corified one at the same PMI. Therefore, histological analysis of elastic fibers in mummified and corified skin may help to provide valuable aid in estimating PMI, especially in those particular cases where more reliable alternatives are lacking.

Utility and diagnostic value of postmortem microbiology associated with histology for forensic purposes.

Nowadays, the diagnostic value of postmortem microbiological investigations is still a debated topic, but postmortem microbiology (PMM) remains a discipline with great forensic potential. To evaluate the usefulness and diagnostic-forensic value of postmortem microbiological cultures, it has been conducted a study on cadaveric material sampled during autopsy aiming to identify the correct cause of death. The study analyzed 45 cadavers subjected to judicial autopsy, divided into two groups based on the presence or absence of external or internal macroscopic autopsy signs suggesting infectious pathology. In the same cases, both the microbiological and conventional histological investigations have been simultaneously carried out. From the investigations, mono-bacterial, mono-fungal, mixed and negative cultures were observed. In mono-species microbiological growth, the histological epicrisis confirmed an infectious cause of death due to the presence of signs of acute infection with an aggressive infectious agent. In cases where growth was mixed, it was possible to distinguish between simple postmortal contamination and perimortem colonization. Finally, in some cases where the microbiology was negative, this has been essential in highlighting signs of a vital reaction to viral or parasitic infection. The joint and integrated evaluation of the laboratory results made it possible to correctly understand even those peculiar situations in which the PMM results alone would not have been significant. These methods, when combined, constitute an optimal forensic approach for the identification of the real cause of death and thus reduce the number of unsolved cases.

Visceral Microscopic Pattern From Suicidal Ingestion of Professional Lysoform® With Delayed Death.

Lysoform® in the formulation of professional detergent is widely used in several fields, whereas its suicidal ingestion is an unusual occurrence. Therefore, the biological signs of this fatal poisoning remain unclear and elusive, similarly to the histological lesions induced by its main constituent, which is benzalkonium chloride (BZK). Furthermore, since all the deaths that has been reported in the literature occurred immediately, microscopic pictures of BZK lethal toxicity in subjects with prolonged survival have never been reported to date. Specifically, this brief communication reports the unique case of a woman who ingested professional Lysoform® to commit suicide, for which she died two weeks later. The autopsy examination showed either local or systemic signs of caustic ingestion; moreover, the histological analysis showed clear cellular damage of lungs, heart, and kidneys. In our case, toxicological investigations were not authorized as they were no longer considered significant. In this framework, the histological examination has therefore assumed a fundamental role in investigating and demonstrating the lethal effects caused by the systemic dissemination of BZK, which would otherwise no longer be investigable. Therefore, in cases of substance intoxication with prolonged survival where forensic toxicological investigations may be no longer possible or feasible, the histological examination may be the only resource to successfully observe and demonstrate its lethal effects.

Postmortem Diagnosis of Invasive Disseminated Aspergillosis After Tongue Piercing.

ABSTRACT: Severe fungal infections caused by highly invasive fungi such as Aspergillus are not easy to diagnose and often have a poor prognosis. In these cases, the nonspecific symptoms may make clinical diagnosis challenging, and consequently, the autopsy and postmortem histological investigations acquire a crucial role. We report the case of a young man in good health who died of septic shock 3 weeks after having had a tongue piercing. Intravitam investigations did not identify the etiology of the rapidly fatal infectious condition. The autopsy revealed flaccid organs of uniformly diminished consistency with abscesses and granulomatous foci with central necrosis. Histological examination showed the presence of septate mycotic hyphae, with a dichotomous 45-degree bifurcation, typical for Aspergillus , in all the examined organs, including the tongue. The molecular identification confirmed the presence of Aspergillus fumigatus. The observed macroscopic framework and the laboratory findings made it possible to diagnose pseudomembranous invasive tracheobronchial aspergillosis and to attribute the death to fatal invasive disseminated aspergillosis. The consistency and concordance of all the findings in our possession led us to suspect the practice of piercing as the triggering cause of the man's pathology.

Technical note: A comparison between rehydrating solutions in the pretreatment of mummified and corified skin for forensic microscopic examination.

Microscopic examination of mummified or corified skin may be of extreme importance for forensic purposes. However, standard histological samples in these cases are low-end, and preparation is burdened by several problems and so are diagnostic results: an improvement of these types of specimens is therefore advantageous. This study aims to identify the best performing rehydration solution among a fabric softener, a body lotion, and Sandison's rehydrating solution. Samples of skin undergoing mummification or corification were collected from 25 corpses and each sample was divided into 4 fragments: one of these fragments was directly fixated in 4% formalin, one was previously treated with a tissue softener, another one was previously treated with a body lotion, and the last one was treated with Sandison's solution. After 72 h, the pretreated samples were post-fixated in 4% formalin and then prepared for standard histological examination staining the histological slides with hematoxylin-eosin and Masson's trichrome. At the microscopic examination, samples directly fixated in formalin were characterized by usual marked structural alterations and altered stainability, typical of such dry tissues. Vice versa, those previously treated appeared to be better-preserved even though with different improvement levels: body lotion made a medium-low-grade restoration of the tissues, and fabric softener a high-grade restoration, while Sandison's rehydrating solution produced an optimal grade restoration. Sandison's rehydrating solution was confirmed to be the best rehydrating substance for mummified and corified skin. Fabric softener could be, however, considered a valid substitute, being productive of high-grade microscopic yield.

Histological microaggregates as a sign of systemic dissemination due to oral ingestion of povidone-iodine (Betadine®).

Povidone-iodine (PVP-I) in the formulation of Betadine® is widely used in several medical fields, whereas its suicidal ingestion is an unusual occurrence. Therefore, the biological signs of this fatal poisoning remain unclear and elusive, similarly to the histological lesions induced by PVP-I. In fact, there are no forensic articles concerning the histological signs of PVP-I acute poisoning. Specifically, this short communication reports the unique case of a man who ingested 125 ml of Betadine® 10% to commit suicide, for which he died three days later. The autopsy examination showed either local or systemic signs of caustic ingestion; moreover, the histological analysis showed both intra- and extra-cytoplasmic amorphous and brownish microaggregates in almost all the organs. Histological and histochemical techniques resulted to be negative for the detection of physiological and known pigments (e.g., formalin, lipofuscin, hemosiderin). Likewise, such pigments were not associated neither with the man's diseases nor with his medications. Therefore, the authors supposed that they were povidone-iodine microaggregates, which have been deposited in the organs through the blood circulation. In conclusion, the complete postmortem histological examination could allow the identification and the characterization of PVP-I microaggregates as evidence of systemic toxicity from Betadine®, when dealing with a forensic case of ingestion of povidone-iodine.

Use of cadaveric vitreous humor as an innovative substrate for diatoms research and forensic diagnosis of drowning.

The search for diatoms is the test that most of all may contribute to the forensic diagnosis of drowning. Diatoms can be extracted through different methods, which are all complex and long. In this preliminary study, we assessed human vitreous humor as an innovative substrate on which to research diatoms. Sampling and analyses were performed on 3 groups of 10 corpses each. The first one was composed of drowned victims. The second one of victims of traffic accidents with subsequent projection in water; based on the severe traumatic injuries reported at autopsy, we considered such victims as ideally assimilable to bodies immersed in water postmortem. The third group was composed of subjects who died for natural causes and without any relationship with water. The vitreous humor was centrifugated and cytocentrifuged and spotted on two different histological slides: one was left blank and the other one was assessed with hematoxylin and eosin staining. Microscopic observation successfully revealed diatoms in all cases of the first group; in the second group diatoms were not detected in 6 cases; in the third group, no diatoms at all were observed. Diatoms were always qualitatively and quantitatively consistent with those found in the corresponding viscera and waters, which had been investigated through classical acid digestion. Our analyses have demonstrated that the vitreous humor behaves in a completely similar way to the other viscera in cases of drowning. Although further investigations are necessary, vitreous humor has proved to be an innovative, suitable, and reliable substrate for the forensic research of diatoms.

Histotopography of haemorrhagic infiltration in the hanging cutaneous furrow: Where to look for haemorrhagic infiltration in hanging.

The forensic evidence of hanging is based on the autopsy demonstration of the traces left by the noose or the ligature on the neck, as well as on the histological assessment of the hanging mark vitality. However, the specific topography of haemorrhagic infiltration in the context of the cervical damaged tissues involved in hanging is not known. We carried out an extensive microscopic examination to identify if haemorrhagic infiltration in hanging appeared in specific topographic locations that could have been considered as elective. From 102 victims of suicidal hanging, a skin fragment was sampled from the maximum compression area of the sulcus, including the skin portions immediately above and below it. The sampling was also extended to collect the subcutaneous adipose tissue and part of the striated muscle. A standard histological examination was performed on all the samples, and hematoxylin and Eosin, Weigert's resorcin-fuchsin and Goldner's Masson trichrome staining were performed. In all the cases assessed, the microscopic examination allowed the detection of haemorrhagic infiltration mainly in the deepest areas under the sulcus and especially in the deep dermis and at the transition point between the dermis and the subcutaneous adipose tissue, as well as in the context of its supporting connective tissue. Such areas could therefore be considered as regions in which the presence of haemorrhagic infiltration is more likely to be demonstrated. Accordingly, we recommend performing sampling similar to ours and focusing the search for haemorrhagic infiltration as suggested.

Analysis of the corrosive effects of hydrochloric acid (HCl) on human bone: Preliminary microscopic study and observations for forensic purposes.

The destruction of a corpse in caustic acid can cause complex forensic scenarios to deal with. Furthermore, the literature on the subject is poor, having been the few studies carried out only on animal bones. We carried out an experimental analysis on human cranial, ribs, vertebrae, and femur bone fragments. These samples were sent for dissolution by hydrochloric acid (HCl) at two different concentrations in the lab: 10% and 37%. We have performed macroscopic and microscopic histological and cytological observations at set time intervals: 3, 4, 19, 24, 48, and 72 h of immersion in acids. The purpose of the study was threefold: to investigate the temporal evolution of bone dissolution, evaluate the destructive effect of the two hydrochloric acids, and establish whether or not the human pattern of histological structure could be recognized. A more significant destructive action of HCl at 37% has been observed. In the 10% acid, the bone nature of the samples was demonstrable up to 24 h of immersion, but the human pattern of histological structure was already compromised at 19 h, being lost at 24 h. Instead, in the 37% acid, the bone nature of the sample was demonstrable only within 4 h of immersion, and the human pattern of histological structure was markedly compromised within 3 h. At 19 h of immersion, neither the recognition of the bone nor its human nature was feasible. These preliminary findings and observations may be of practical use in forensic investigations of bodies found in acidic substances, for which there is no scientific evidence to refer.

Paradoxal dyeing affinity's inversion of the connective tissue at Goldner's Masson trichrome staining as a peculiar characteristic of compressed and exsiccated cadaveric skin.

The microscopic examination of a hanging cutaneous furrow, stained with Goldner's Masson trichrome staining, highlighted an abnormal dyeing inversion affinity of the connective tissue - red instead of green - located in the region of maximal cutaneous compression and exsiccation. To identify if this different stainability could be considered as an intrinsic characteristic of all biological tissues compressed and exsiccated, we have assessed different cadaveric skin samples that underwent traumatic detrimental phenomena that can produce such effects. We collected skin fragments from 24 corpses deceased because of gunshot injuries, electrocution, hanging, and heat-induced lesions, sampling the areas directly involved, as well as skin specimens to use as a control. The slides were stained with hematoxylin and eosin and two different protocols of Goldner's Masson trichrome staining (one homemade and one commercial kit). The inversion of the staining affinity of the connective tissue was observed in 83% of the cutaneous samples, using both the Goldner's Masson trichrome staining protocols. This phenomenon was not observed in any of the control cases. Therefore, the inversion of the staining affinity of the connective tissue dyed with Goldner's Masson trichrome staining may represent a histomorphological aspect that must be expected when the skin has been affected by specific detrimental modalities able of producing compression and exsiccation.

Lethal extracranial hemorrhage due to intracranial clivalchordoma: An autopsy case.

We present a case of a 48-year-old man's unexpected death affected by a relapsed clivalchordoma. After partial excision surgery of the neoplasm, he manifested 5 days later, in conditions of well-being, a sudden lethal extracranial hemorrhage from nose and mouth. The autopsy examination and the subsequent histological investigations did not allow us to clarify the exact origin of the bleeding. Based on the negativity of the accurate examinations performed, the extent of the bleeding, and the findings highlighted by the means of the nuclear magnetic resonance (NMR) carried out a few days before death, we have considered reasonable to localize the source of hemorrhage in the intrapetrous tract of the left internal carotid artery. Since this is a unique event, never previously documented, we believe that our report may be of interest to the scientific community.

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.