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.

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.

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.

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.

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.

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.

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.

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.

Repeated attempted homicide by administration of drugs documented by hair analysis.

Attempted murder by repeated poisoning is quite rare. The authors describe the case of a 62-year-old man who was admitted to an intensive care unit (ICU) for neurological disturbances complicated by inhalation pneumopathy. He presented a loss of consciousness while his wife was visiting him at the ICU (H0). Forty-eight hours later (H48), police officers apprehended the patient's wife pouring a liquid into his fruit salad at the hospital. Toxicological analyses of a blood sample and the infusion equipment (H0), as well as the fruit salad and its container (H48), confirmed the attempted poisoning with cyamemazine (H0) and hydrochloric acid (H48). In order to evaluate the anteriority of poisonings, hair analysis was requested and the medical records of the 6 previous months were also examined. Two 6-cm brown hair strands were sampled and the victim's medical record was seized in order to determine the treatments he had been given during the previous six months. Segmental hair testing on two 6-cm brown hair was conducted by GC-MS, LC-DAD and LC-MS/MS (0-2/2-4/4-6 cm; pg/mg). Haloperidol (9200/1391/227), amitriptyline (7450/1850/3260), venlafaxine (332/560/260), that had never been part of the victim's treatment were detected, as well as some benzodiazepines (alprazolam, bromazepam, nordazepam); cyamemazine was also detected in all the segments (9960/1610/2367) though only a single dose administration was reported in the medical records. The toxicological analyses performed at H0 and H48 confirmed the homicide attempts in the ICU. In addition, comparison of the results in hair analysis with the medical records confirmed repeated poisoning attempts over the previous six months, and thus explain the origin of the disorders presented by the victim. This case serves to remind us that repeated attempted murder can be difficult to diagnose and that hair analysis can be an effective way to detect such attempts.

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.

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.

Post Mortem Redistribution of Drugs: Current State of Knowledge.

BACKGROUND: Drug concentrations obtained from post mortem samples do not necessarily reflect the concentrations at the time of death, and variations of concentration may be observed between different sites and/or different sampling times. These phenomena, collectively termed post mortem redistribution, concern numerous molecules (medications, drugs of abuse, gases, etc.) and can complicate the interpretation of toxicological analyses. METHODS: Literature review. RESULTS: The mechanisms that cause these phenomena are complex and often intricate. Certain organs, which concentrate the molecules before death, may release them very early in the vascular sector. The gastrointestinal tract, liver, lungs and myocardium are mainly concerned. Cell autolysis also plays a part in drug release. Furthermore, micro-organisms (mainly bacteria and yeasts) which colonize the organism during putrefaction may cause neoformation and/or the degradation of certain molecules. Lastly, it appears that the physicochemical and pharmacokinetic profile of xenobiotics, notably their lipophilic nature, their ionization state and their volume of distribution may be factors likely to influence redistribution phenomena. Some recommendations concerning anatomic sampling sites, sampling methods and sample storage make it possible to limit these phenomena.

Medicolegal Implications of Nosocomial Infections: A Case Report of Aspergillus Contamination during Cardiac Surgery.

Nosocomial infections have become a major issue of public health and lead to an increasing number of suits for damages. We present a rare case of Aspergillus contamination during cardiac surgery, describe the medicolegal investigation, and present the new system for compensation of bodily injury after nosocomial infection in France, based on the law of March 4, 2002 on patient rights and quality in the health system. This case demonstrates the limits of compensation for nosocomial infections on the grounds of national solidarity. The expert report requested by the regional commission for conciliation and compensation is of fundamental importance in enabling the commission to decide between fault and inherent risk of treatment.

Death from Hypothermia during a Training Course under "Extreme Conditions": Related to Two Cases.

Death from hypothermia following exhaustion or from various complicated pathologies is no longer a frequent cause of death among combat troops. During a training course under "extreme conditions" in the French Alps, two young African officers died. Confronted with these two clinically confirmed cases of hypothermia, the unknown anatomopathological and biological specificities associated with death from hypothermia were highlighted. In these typical and clinically confirmed cases of death from subacute exhaustion hypothermia, none of the signs revealed by the autopsy were specific. Although some recent publications have addressed the utility of postmortem biochemical markers when establishing a diagnosis, with no anamnesis, with no knowledge or analysis of the circumstances of death, and without an in situ examination of the body, it appears difficult, if not impossible, to confirm that death was caused by hypothermia.

Transmission of force to the hyoid bone during manual strangulation: Simulation using finite element numerical models.

INTRODUCTION: Strangulation is a medicolegal form of mechanical asphyxia, and can be difficult to identify if cutaneous damage to the neck is limited. We began by creating a numerical model of a hyoid bone with adjustable anthropometric parameters and then subjected our model to compression simulating a precision grip on the neck from the front. MATERIALS AND METHODS: We selected six bones from the 77 hyoid bones contained in the database we created during a previous study led by our laboratory, in which we demonstrated the sexual dimorphism of this bone and identified the minimal force required for fracture. The anthropometric characteristics of these six bones (angle, length and width) corresponded to those of the 10th, 50th and 90th percentiles from cluster 1 (male individuals) and cluster 2 (female individuals), respectively. After enhancing, developing and meshing the selected 3D models, we analysed and reproduced simulation conditions that were as close as possible to the in vivo conditions of neck strangulation from the front (area of pressure on the bone, tissue environment, and biological variability of this bone). RESULTS: We modelled the six numerical hyoid bone models using the finite element method. For all models, the simulation of mechanical pressure applied to simulate anterior strangulation resulted in fractures. The forces required to produce these fractures matched the results obtained in the experimental testing of dissected hyoid bones. CONCLUSION: Six finite element numerical models were created, covering the anthropometric morphological variability of the hyoid bone. These six models enabled numerical simulation of the in vivo behaviour of a hyoid bone subjected to one-handed strangulation.

Hair testing in postmortem diagnosis of substance abuse: An unusual case of slow-release oral morphine abuse in an adolescent.

Morphine sulfate misuse is essentially observed among regular heroin injectors. To our knowledge, primary addiction to morphine sulfate is exceptional, especially among young adolescents. A 13-year-old girl, with no history of addiction, was found dead with three empty blisters of Skenan(®) LP 30 mg at her side. Opiates were detected in biological fluids and hair by chromatographic methods. Blood analyses confirmed morphine overdose (free morphine: 428 ng/mL; total morphine: 584 ng/mL) and segmental hair analysis confirmed regular exposure over several months (maximum morphine concentration 250 pg/mg). Suspecting the victim's mother of recreational use of Skenan(®), the magistrate ordered analysis of her hair, with negative results. From an epidemiological viewpoint, this case of oral morphine sulfate abuse in an adolescent with no previous history suggests the emergence of a new trend of morphine sulfate consumption. From a toxicological viewpoint, it demonstrates the value of hair testing, which documented the victim's regular exposure and made an important contribution to the police investigation.