Histological dating of subdural hematoma in infants.

BACKGROUND: After infant deaths due to non-accidental head injury (NAHI) with subdural hematoma (SDH), the magistrates ask experts to date the traumatic event. To do so, the expert only has tools based on adult series of NAHI. We aimed to develop an SDH dating system applicable to infants aged under 3 years. METHODS AND RESULTS: We studied a retrospective multicenter collection of 235 infants who died between the ages of 0 and 36 months, diagnosed with SDH by forensic pathological examination and with known posttraumatic interval (PTI). Two pathologists assessed blindly and independently 12 histomorphological criteria relating to the clot and 14 relating to the dura mater in 73 victims (31 girls, 42 boys) whose median age was 3.8 months. Histopathological changes were significantly correlated with PTI for the appearance of red blood cells (RBCs) and the presence or absence of siderophages, and regarding the dura mater, the quantity of lymphocytes, macrophages, and siderophages; presence or absence of hematoidin deposits; collagen and fibroblast formation; neomembrane thickness; and presence or absence of neovascularization. Dating systems for SDH in adults are not applicable to infants. Notably, neomembrane of organized connective tissue is formed earlier in infants than in adults. CONCLUSION: Our dating system improves the precision and reliability of forensic pathological expert examination of NAHI, particularly for age estimation of SDH in infants. However, the expert can only define a time interval. Histopathology is indispensable to detect repetitive trauma.

High-Throughput Monitoring of Cocaine and Its Metabolites in Hair Using Microarrays for Mass Spectrometry and Matrix-Assisted Laser Desorption/Ionization-Tandem Mass Spectrometry.

Because of inhomogeneous matrix-assisted laser desorption/ionization (MALDI) matrix crystallization and laser shot-to-shot variability, quantitation is not generally performed by MALDI mass spectrometry. Here we introduce a high-throughput MALDI method using an innovative high-density microarray for mass spectrometry (MAMS) technology, which allows semiquantitative measurement of cocaine and its metabolites, benzoylecgonine, cocaethylene, and ecgonine methyl ester. A MAMS slide containing lanes of hydrophilic spots and an automated slider to drag a sample droplet over several small spots can accomplish automatic sample aliquoting and lead to homogeneous crystallization of the matrix-analyte mixture and, thus, to a reproducible signal (average RSD 6%). Four hair samples of self-reported drug users were analyzed in parallel by MALDI-MS/MS and by a validated LC-MS/MS method. The consumption profiles as well as the metabolite-parent drug ratios obtained correlated well, confirming the effectiveness of the MALDI-MS/MS method to establish a calendar of consumption in only 1 mg of hair. The analysis time for 10 hair samples is below 40 min, with 12 replicates per sample. Since only 3 µL of a 20 µL extract is analyzed, complementary assays are possible, such as the detection of additional drugs. The semiquantitative MALDI method worked well with only a small amount of hair and gave results in less than 4 min per sample, including replicates. This was made possible by the use of MAMS slides for sample preparation, which thus present significant advantages over traditional methods in cases where results are required urgently or if samples are scarce.

Age determination of adult individuals by three-dimensional modelling of canines.

Accurate age determination is fundamental in both forensic medicine and anthropology. Many methods that relate dental characteristics to adult age have been proposed, but there is still no simple and reliable method that does not damage the study material. The aim of this work was to propose a relevant and practical technique for determining age in adults that could be used in both living and deceased individuals. The sample was composed of 210 CT scans from individuals aged from 15 to 85 years old, with four healthy canines present in the mouth. The 840 canines were modelled using Mimics® 10.01 software. The pulp volume/total volume ratio ×100 was determined for each tooth. Seven mathematical models, corresponding to all possible real situations, were determined by the weighted least squares method and ranked in order of relative performance. The adequacy of the seven models to the data was very high with the regressions proposed (0.915 < R (2) adjusted < 0.964). Ranked in order of performance, the maxillary model was the most powerful of the seven models for age determination, followed by the 4 canines model, the 13 model and the 23 model.

Hair analysis in postmortem investigations: Case of a skeletonized body.

Analysis of hair collected from putrefied or skeletal bodies is always complex and must take into account several pitfalls, such as external contamination and contamination by biological fluids. This work presents a case of particular complexity. A skeletonized body was discovered on a country road. A tuft of brown hair, detached from the scalp, irregular in length, non-oriented, in contact with soil and vegetation, was removed. An anthropological examination was carried out and genetic samples were taken from the right femoral shaft. After about 10 washes with warm water and dichloromethane, the tuft of hair was analyzed without segmentation. General unknown screening was performed by liquid chromatography system coupled to a high-resolution mass spectrometry (LC-HRMS) after incubation in pH 9.5 borate buffer and liquid-liquid extraction. Specific Multiple Reaction Monitoring (MRM) methods for date rape drugs were carried out by liquid chromatography system coupled to a tandem mass spectrometry (LC-MS/MS). The anthropological examination allowed to determine that the victim was a female individual, over 60 years old, the death dating from 3 months to 1 year. Comparison of the DNA results with the Missing Persons Index led to the identification, a 60-year-old woman who disappeared 5 months earlier. Hair analysis showed the presence of oxazepam (361 pg/mg), nordiazepam (54 pg/mg), and alimemazine (5 pg/mg). The interpretation of these concentrations is extremely difficult due to the risk of degradation of the hair cuticle during prolonged stay in the soil, as well as of contamination by putrefactive fluids. The authors discuss the value of using multiple biological and non-biological matrices in this context to improve the interpretation of the results.

Effects of fall conditions and biological variability on the mechanism of skull fractures caused by falls.

In a forensic investigation, there is considerable difficulty in distinguishing between different mechanisms that could explain the head injury sustained. The key question is often whether the injury was the consequence of a fall, a blow, or a fall caused by a blow. Better understanding of the parameters influencing the mechanism of skull fracture could be of use when attempting to distinguish between different causes of injury. Numerous parameters concerning fall conditions and biological variability are reported in the literature to influence the mechanism of skull fracture. At the current time, there are no studies that investigate both the effect of a fall and biological parameters. The aim of this paper is to study the influence of these parameters on the mechanism of skull fracture using a numerical approach. We focused on accidental falls from a standing height. A multibody model was used to estimate head impact velocities and a finite element model was used to investigate the effect of the fall conditions and of biological variability on skull fracture. The results show that the mechanism of skull fractures is influenced by a combination of at least four parameters: impact velocity, impact surface, cortical thickness and cortical density.

Can subclavian blood be equated with a peripheral blood sample? A series of 50 cases.

If femoral blood is not available at autopsy, toxicological analyses, in particular blood ethanol measurements, are carried out on cardiac blood. This is known to be subject to major redistribution. We aimed to determine whether subclavian blood can be equated with a peripheral blood sample and could be used if femoral blood is not available. The study was based on 50 medicolegal autopsies in which we compared ethanol concentrations between subclavian blood, the different heart blood compartments (right and left cardiac blood), and femoral blood. Mechanisms that could lead to variations in concentration, i.e., postmortem redistribution and/or endogenous production, were also taken into account in interpreting the results. Ethanol concentrations were determined by headspace gas chromatography with a flame ionization detector. In each case, we recorded the circumstances of death, resuscitation attempts if any, degree of putrefaction, chest or abdominal trauma, and/or inhalation of gastric fluid in the airways. Ethanol concentrations in subclavian blood were found to be close to those in peripheral blood (p = 0.948) and were not influenced by the degree of putrefaction (r = 0.017, p = 0.904), gastric ethanol concentration (r = -0.011, p = 0.940), inhalation of gastric contents in the airways (p = 0.461), or cardiac resuscitation attempts (p = 0.368). We discuss the possible explanations for these findings and stress the value of sampling subclavian blood when femoral blood is not obtainable at autopsy.

Recent Trends in MALDI-MS Drugs Analysis in Human Hair: Pre-Analytical and Analytical Challenges and Pitfalls.

The value of hair in forensic toxicology has already been widely demonstrated. It offers a much wider detection window than other matrices, and its segmental analysis allows the documentation of a single, occasional or regular consumption of a large number of molecules. To date, considerable efforts are being made to achieve very high sensitivity using increasingly effective techniques in the forensic analysis of hair (gas chromatography with electron ionization mass spectrometry (GC--EI-MS); gas chromatography with chemical ionization mass spectrometry (GC--CI-MS); gas chromatography-tandem mass spectrometry (GC--MS-MS); high or ultra performance liquid chromatography-tandem mass spectrometry (HPLC-MS-MS and UPLC-MS-MS)). Since the early 2000s, work has been carried out to analyze hair using matrix-assisted laser desorption ionization (MALDI) coupled with imaging mass spectrometry (IMS). Intact, cut or pulverized, human head hairs are analyzed in all their forms. With a simplified and rapid sample preparation protocol, MALDI-IMS appears to be an attractive option for the forensic interpretation of hair analysis. The high spatial resolution clearly competes with conventional methods and strand segmentation. This article provides a complete overview on MALDI techniques used and its applications to better understand the pre-analytical and the analytical parts in the hair analysis.

Penetrating craniocerebral injury caused by bladed weapons: Three homicide cases.

Penetrating craniocerebral injuries caused by a bladed weapon are rare events in western countries and often occur in the course of assault. We studied all homicide-related cases of cranial and brain injuries caused by bladed weapons that were seen at the Medicolegal Institute of Marseille over a 5-year period from 2014 to 2019. We found that such injuries are exceptional. The majority occur in the private sphere and are inflicted by men, as they require considerable force. They are rarely isolated but are accompanied by multiple cutting injuries of the thorax and abdomen. Such assaults leave imprints on the bone that can be analyzed using new anthropological techniques.

[A rare cause of sudden cardiac failure: histiocytoid cardiomyopathy]. / Une cause rare de défaillance cardiaque brutale chez le nourrisson : la cardiomyopathie histiocytoïde.

Histiocytoid cardiomyopathy is a rare disease which occurs predominantly in the first two years of life, with a female preponderance. We report the cases of two girls (11 and 15-month-old) which were respectively referred to our institution for ventricular tachycardia and ventricular fibrillation without prodroma. Etiologic findings only showed mild cardiomyopathy. Autopsy and histologic examination led to the diagnosis of histiocytoid cardiomyopathy. Furthermore, in the first observation, agenesis of the corpus callosum was found.

Planned complex suicide involving combined drug intoxication and femoral catheterization.

Complex planned suicide is characterized by the simultaneous use of two or more methods to ensure that death occurs even if one method fails. The authors present an original combination of two self-killing methods. A 42-year-old cardiologist, with a major depressive syndrome and several suicide attempts, as well as cocaine addiction, was found dead at his home with a femoral catheter inserted in the right femoral artery. The autopsy concluded that death was due to major hemorrhagic process in a context of suicide. Toxicological analyses, performed in peripheral blood by gas chromatography coupled to mass spectrometry and by liquid chromatography-diode array detection, revealed the presence of ethanol (0.13 g/L), cocaine, and metabolites (cocaine: 432 µg/L, benzoylecgonine: 3286 µg/L, ecgonine methyl ester: 1195 µg/L, cocaethylene: 41 µg/L), a potentially lethal concentration of citalopram (1.03 mg/L), toxic concentrations of hydroxyzine (0.11 mg/L), bromazepam (2.06 mg/L), and lidocaine (7.30 mg/L). At the end of these analyses, the death was reclassified as planned complex suicide combining drug intoxication and catheterization of the femoral artery. The authors discuss the main aspects of this case and stress the importance of meticulous analysis of all available evidence: witness reports, victim's medical history and occupation, findings of at-the-scene examination, autopsy, and toxicological analyses, in order to exclude homicide and to understand the sequence of events that led to death.

Homicides and suicides by firearm in Marseille: An 8-year review.

The regions of Provence-Alpes-Côte d'Azur and Corsica in south-east France are regularly affected by firearm deaths, and a large amount of data has been collected by our forensic institute. We carried out a single-center retrospective descriptive study of the records of our institute between January 1, 2011, and 31 December, 2018, relating to firearm deaths (homicides and suicides). There were 302 cases (218 homicides and 84 suicides). The anatomic locations most frequently involved were the thorax, abdomen and head, in that order, in homicides and the head in suicides. More than 80% of the homicides in our series had more than one wound complex and nearly one in four homicides had more than 7. The weapon most frequently used in suicides was a hunting smooth bore gun, followed by the 9mm Luger and then by the .22 Long Rifle. In homicides, the caliber most frequently used was the 7.62 × 39 (29.6%), followed by the 9mm Luger (29%) and lastly by hunting calibers (27%). More than 10% of cases involved two weapons of two different calibers. Our study, which used a dual forensic and ballistic approach, was carried out in one of the most heavily populated regions of metropolitan France. We observed very few similarities with the literature regarding type of caliber and wound complexes. This may be explained, among other factors, by differences in political, societal or cultural contexts.

Buprenorphine-related deaths: unusual forensic situations.

The success of high-dose buprenorphine (HDB) as substitution therapy for major opioid dependence is related to its partial agonist effect on opioid receptors, which in theory makes it very safe to use. However, numerous deaths directly attributable to buprenorphine have been described in the literature. These deaths are generally related to misuse of HDB with intravenous administration and/or concomitant use of benzodiazepines, and they usually occur in patients on HDB substitution therapy for opioid dependence. We present three deaths attributed to HDB which arose from uncommon mechanisms and led to unusual forensic situations. The first death was that of a patient admitted to hospital after simultaneous prescription of HDB, clonazepam, oxazepam, and cyamemazine. The second death followed forcible administration of a very low dose of HDB to a patient with post-hepatitis C cirrhosis and heart failure. The third death was subsequent to an HDB overdose, probably with suicidal intent, in a young woman who had not been prescribed the drug as opiate substitute. Such deaths raise the question of the mechanisms involved and draw attention to the resulting unusual forensic situations.

Lethal injection of potassium chloride: first description of the pathological appearance of organs.

Lethal injection of potassium chloride (KCl) can be used as a method of either suicide or homicide. As biological tests are still inadequate to differentiate endogenous from exogenous potassium, at the scene of death the cause can only be suspected. We wished to determine the usefulness of conventional pathological examination in this context and carried out a study in four fetuses after medical termination of pregnancy for serious disease. Pregnancy was terminated by KCl injection in two cases and by injection of lidocaine and sufentanil in the other two cases. In each of the two fetuses in which KCl injection was performed, macroscopic examination showed whitish deposits on the tissues and histological examination showed clumps of lanceolate crystals in the internal organs. In the two fetuses which received lidocaine and sufentanil injection, no deposits were visible on macroscopic examination and no crystals were seen on histological examination. These findings suggest that pathological study may have useful applications in forensic medicine when death by potassium injection is suspected.

Study of cerebrospinal injuries by force transmission secondary to mandibular impacts using a finite element model.

Brain and cervical injuries are often described after major facial impacts but rarely after low-intensity mandibular impacts. Force transmission to the brain and spinal cord from a mandibular impact such as a punch was evaluated by the creation and validation of a complete finite element model of the head and neck. Anteroposterior uppercut impacts on the jaw were associated with considerable extension and strong stresses at the junction of the brainstem and spinal cord. Hook punch impacts transmitted forces directly to the brainstem and the spinal cord without extension of the spinal cord. Deaths after this type of blow with no observed histological lesions may be related to excessive stressing of the brainstem, through which pass the sensory-motor pathways and the vagus nerve and which is the regulatory center of the major vegetative functions. Biological parameters are different in each individual, and by using digital modeling they can be modulated at will (jaw shape, dentition…) for a realistic approach to forensic applications.

Using an ultraviolet cabinet improves compliance with the World Health Organization's hand hygiene recommendations by undergraduate medical students: a randomized controlled trial.

BACKGROUND: Appropriate hand hygiene (HH) is key to reducing healthcare-acquired infections. The World Health Organization (WHO) recommends education and training to improve HH knowledge and compliance. Physicians are ranked among the worst of all healthcare workers for compliant handrubbing with its origin probably being the failure to learn this essential behavior during undergraduate medical studies. This study evaluated if the use of Ultraviolet-cabinets (UVc) for fluorescent-alcohol-based handrubs (AHR) during an undergraduate medical student training improved the compliance rate to the WHO hand hygiene recommendations (completeness of AHR application and HH opportunities). METHODS: This randomized trial compared a HH training with personal feedback (using UVc) to a control group. The first year, the students (2nd degree) were convened by groups (clusters) of 6-9 for a demonstration of the correct execution of WHO procedure. Randomization by cluster was done prior HH training. In the control group, the students hand rubbed under visual supervision of a tutor. In the intervention group after the same visual supervision, completeness of fluorescent-AHR hand application was recorded under UVc and was shown to the student. The intervention group had free access to the UVc until complete application. HH practices were included in simulation sessions for the both groups. One year after (3rd degree), all the students were asked to hand rub with fluorescent-AHR. A tutor (blinded to the study group) assessed the completeness of hand application under UVc and the compliance with the WHO opportunities. Complete application of AHR was defined as fluorescence for all the surfaces of hands and wrists. RESULTS: 242 students participated (140 in the intervention group and 102 in the control group). One year after the initial training, the rate of complete application of AHR was doubled in the intervention group (60.0% vs. 30.4%, p < 0.001). In a multivariate analysis which included gender, additional HH or UVc training, surgical traineeship and regular use of AHR, the hazard ratio for the intervention was 3.84 (95%CI: 2.09-7.06). The compliance with the HH WHO's opportunities was increased in the intervention group (58.1% vs. 42.4%, p < 0.018). CONCLUSION: Using UVc for undergraduate medical students education to hand hygiene improves their technique and compliance with WHO recommendations.

Histological dating of subarachnoid hemorrhage and retinal hemorrhage in infants.

BACKGROUND: Dating the traumatic event is usually done on subdural hematoma (SDH). After infant deaths due to Abusive head trauma (AHT) without SDH available, the magistrates still ask experts to date the traumatic event. To do so, the expert only has tools based on adult series of AHT. We aimed to develop a subarachnoid hemorrhage (SAH) and retinal hemorrhage (RH) dating system applicable to infants aged under 3 years. METHODS AND RESULTS: We studied a retrospective multicenter collection of 235 infants who died between the ages of 0 and 36 months, diagnosed with SAH and/or RH by forensic pathological examination and with known posttraumatic interval (PTI). Two pathologists assessed blindly and independently 12histomorphological features in 83 infants (35 girls, 48 boys) whose median age was 3.8 months. For SAH, histopathological changes were significantly correlated with PTI for the appearance of red blood cells, of fibrino-plaquetted organization, the quantity of lymphocytes and macrophages and the presence or absence of siderophages, collagen and fibroblast formation and presence or absence of neovascularization. For RH, histopathological changes were significantly correlated with PTI for the appearance of red blood cells, the presence or absence of siderophages and sclerosis of the retina. CONCLUSION: Our HAS dating system improves the precision and reliability of forensic pathological expert examination of AHT, when SDH are not available, for age estimation in infants. The study of RH histomorphological changes does not allow for reliable dating.

Sudden unexpected infant death characteristics in the French region of West Provence-Alpes-Côte d'Azur.

Background: Although the incidence of sudden unexpected infant death (SUID) has decreased since the 'Back to Sleep' campaign in English-speaking countries and other preventive campaigns, the circumstances of such deaths remain unclear. Aim: To analyse infant deaths recorded at the referral centre for sudden infant death of the West Provence-Alpes-Côte d'Azur region of France (West PACA) and the forensic medicine department of Marseille University Hospital. Methods: Information on all SUID cases from 2000 to 2017 was extracted from the referral centre for sudden infant deaths in West PACA and the forensic medicine department of Marseille. Results: The study included 130 infants over the 17 years with a very similar distribution. There was a marked male preponderance, with 61.6% of boys whatever the age at death (sex ratio 1.6). Half of the deaths occurred in the first 6 months of life and the majority (61%) of infants died during autumn and winter. Nearly one-third (33.2%) had presented with minor infections and 21% had been seen by a doctor or had been admitted to hospital. Most deaths (86.4%) occurred during sleep (night or day). Nearly half of the infants (47.7%) were discovered in a prone position. A large majority of parents (90.7%) agreed to a post-mortem examination. Only 6.2% of deaths led to legal proceedings. Nearly 16.9% remained unexplained after compiling all the data included in the protocol and 9.2% remained unexplained because of incomplete investigation, including refusal of post-mortem examination. Abuse was involved in 2.3% of cases. Conclusions: Asymptomatic infectious conditions were associated with a high proportion of SUID cases. Non-supine sleep positions were still practised. There is a need to increase SUID prevention campaigns. Abbreviations: HAS, Haute Autorité de Santé: French National Health Authority; NICHD, National Institute of Child Health and Human Development; PACA, Provence-Alpes-Côte d'Azur region of France; SUID, sudden unexpected infant death; SIDS, sudden infant death syndrome; CépiDc, Centre d'Epidémiologie sur les Causes Médicales de Décès/Center for Epidemiology on the Medical Causes of Death.

[A potential contribution of virtual slides and automatic image processing to the estimation of the vascularization of cranial sutures]. / Apport potentiel des lames virtuelles et de l'analyse d'images automatique dans l'estimation de la vascularisation des sutures crâniennes.

AIM: Determining age at the time of death is a difficult problem in forensic practice. The study of the vascularization of cranial sutures is an original approach, which may mark the process of synostosis associated with aging. Counting sections of blood vessels on a histological section of cranial suture, raises however a number of practical problems related to the quality of the preparation and to the method of quantification. The purpose of this study is to illustrate the potential contribution of an automatic analysis of virtual slides to overcome these problems. MATERIALS AND METHODS: The proposed method of analysis is illustrated from three samples of frontosphenoidal suture whose vessels were immunostained after decalcification of bone tissue. A high resolution image (x 20 objective and microscopic resolution of 0.5 microm) of each microscopic preparation was acquired through a microscopic scanning device. The automatic image analysis protocol takes advantage of a processing of virtual slides at two resolutions. RESULTS: The chosen strategy ensures the identification of the specific area of interest, the enumeration of blood vessel sections on the whole preparation and the visual control of the detected structures. CONCLUSION: The quantitative estimate of the vascularization of a large structure, such as a cranial suture, can benefit from scanning and fully automatic processing of virtual slides. The automatic analysis requires however an optimal preparation of tissues; decalcification and immunohistochemical staining must be done with great care.

Suicide by Medication Overdose in Prison: A Study of Three Cases.

Suicide is one of the principal causes of mortality in a prison environment. Although suicide by medication overdose is less frequent than suicide by hanging, self-strangulation, or vein cutting, it raises questions as to how the medications are obtained, particularly in view of the specific organization of the medication circuit in prisons. We present three cases of suicide by medication overdose involving different therapeutic classes with different distribution circuits and review the regulatory requirements and the measures that could be taken to prevent such suicides.

Synthetic cannabinoid isomers characterization by MALDI-MS3 imaging: Application to single scalp hair.

New designer drugs, as synthetic cannabinoids (SCs), continuously appear on the market and are booming due to their cannabis-like effect. New generation of smokable SCs, structurally dissimilar from Δ9-tetrahydrocannabinol (THC), have isomers with distinguishable pharmacokinetic parameters and therefore different in vivo effects. The isoforms are misidentified using conventional techniques such as gas or liquid chromatography coupled to mass spectrometry - or tandem mass - spectrometry. The aim of this study was to differentiate three positional isomers (JWH-007, JWH-019 and JWH-122) in single human hair samples, which store numerous substances revealing a way of life and consumption style. Matrix-assisted laser desorption/ionization (MALDI) combined with imaging is an innovative and powerful tool used since few years, especially in forensic research. Herein, we propose an innovative method to monitor the drugs of abuse consumption through direct mapping of the compounds with a high spatial distribution in human hair samples, by state-of-art imaging MALDI-MSn. Three positional SC isomers (JWH-007, JWH-019 and JWH-122) were analysed using high and low fragmentation energy and the resulting MS/MS and even MS3 spectra differentiated the SCs. The MALDI-MS/MS and MS3 imaging was performed on hair soaked in a mixture of the three SCs as well as on hair from self-reported SC user, proving the potential of the technique for a forensic use. Keeping in mind that spatial distribution of organics from whole hair remains a challenge, the described methodology is a very promising analytical tool to probe the consumption of complex drugs and obtain correlation with its origin.