Current status of undergraduate teaching in forensic & legal medicine in Europe.

The European Council of Legal Medicine (ECLM) is the body established in 1992 to represent practitioners forensic & legal medicine and is composed of delegates of the countries of the European Union (EU) and from other countries which form part of Europe to a current total of 34 member countries. The aims of this study were to determine the current status of undergraduate forensic & legal medicine teaching in the curriculum of medical studies in ECLM countries and to use the results of this study to determine whether it would be appropriate to develop new guidelines and standards for harmonising the content of undergraduate forensic medicine training across ECLM member countries. A detailed questionnaire was sent to all individuals or organisations listed on the ECLM contact database. Responses were received from 21 of 33 countries on the database. These responses showed considerable emphasis on undergraduate teaching of forensic medicine in all countries with the exception of Belgium and the United Kingdom. There was great general consistency in the subjects taught. The data from this survey provide a baseline which should assist in developing a strategy to harmonise forensic & legal medicine undergraduate training in member countries of the ECLM. The ECLM is now in a good position to establish a pan-European working group to coordinate a consensus document identifying an appropriate and modern core undergraduate forensic medicine curriculum that can be presented to the medical education authorities in each country, and which can be adapted for local requirements, based on available personnel, the forensic medicine structure in the country, and most importantly, the needs of the local population.

Be aware of (R)-methamphetamine: Negative immunoassay versus positive confirmation analysis.

The amphetamine-type stimulant methamphetamine exists in two enantiomeric forms, (S)-methamphetamine and (R)-methamphetamine, which are both psychoactive but with the (S)-enantiomer being more potent than the (R)-enantiomer. Illicit methamphetamine encountered in Europe is typically a racemic mixture of both enantiomers and enantiopure (S)-methamphetamine, respectively. However, herein we report two cases with proven enantiopure (R)-methamphetamine consumption with moreover both cases remaining undetected by immunoassay screening. Inconspicuous immunoassay findings can be traced back to a considerably higher sensitivity and concentration-dependent cross-reactivity of the applied drug of abuse assay for the (S)-enantiomer of methamphetamine compared with the (R)-enantiomer, and this limitation should be well known by users of immunoassay drug tests.

Death on New Year's Eve caused by illegal fireworks-a firework shell.

During the turn of the year, injuries caused by fireworks occur in Germany every year. According to the professional associations, the Unfallkrankenhaus Berlin, for example, treats an average of 50 injuries caused by fireworks on New Year's Eve. Patients come with burns, soft tissue injuries, or fractures; eyes and hands are particularly frequently affected. Again and again, there are also very serious or even fatal injuries. The background is usually the improper or illegal use of larger fireworks. Smaller fireworks such as sparklers, bangers, or smaller rockets are available in Germany in most supermarkets, and their use is permitted from the age of 12 or 18. However, the use of larger fireworks in Germany requires proof of an official permit to handle pyrotechnic objects, which is why they are often acquired abroad. The following report describes such a case. Shortly after the turn of the year 2022, a young man died as a result of an explosive effect on the facial skull after using an illegal firework shell. The case is discussed with regard to the autopsy findings, the possible cause of the accident, and the type of firework used.

Cost-Effective Next Generation Sequencing-Based STR Typing with Improved Analysis of Minor, Degraded and Inhibitor-Containing DNA Samples.

Forensic DNA profiles are established by multiplex PCR amplification of a set of highly variable short tandem repeat (STR) loci followed by capillary electrophoresis (CE) as a means to assign alleles to PCR products of differential length. Recently, CE analysis of STR amplicons has been supplemented by high-throughput next generation sequencing (NGS) techniques that are able to detect isoalleles bearing sequence polymorphisms and allow for an improved analysis of degraded DNA. Several such assays have been commercialised and validated for forensic applications. However, these systems are cost-effective only when applied to high numbers of samples. We report here an alternative, cost-efficient shallow-sequence output NGS assay called maSTR assay that, in conjunction with a dedicated bioinformatics pipeline called SNiPSTR, can be implemented with standard NGS instrumentation. In a back-to-back comparison with a CE-based, commercial forensic STR kit, we find that for samples with low DNA content, with mixed DNA from different individuals, or containing PCR inhibitors, the maSTR assay performs equally well, and with degraded DNA is superior to CE-based analysis. Thus, the maSTR assay is a simple, robust and cost-efficient NGS-based STR typing method applicable for human identification in forensic and biomedical contexts.

Histological examination of carotid artery tissue in cases of ligature strangulation and hanging.

Violence against the neck can result in a range of macromorphological and micromorphological findings. However, the forensic relevance of the carotid sinus in cases of violence against the neck remains controversial. In this follow-up study of 22 cases of suicidal and accidental strangulations, carotid bifurcations were examined histologically for morphological changes implying direct trauma, including haemorrhage and immunohistochemical expression of heat-shock proteins 27, 60, and 70 and aquaporin-3. These cases were compared with a control group (82 cases) without neck compression or head trauma and with variable causes of death. No relevant histopathological findings implying direct trauma of the carotid bifurcation were found. No cases showed positive aquaporin-3 staining and only five cases showed positive heat-shock protein-27 staining, all of which were hangings. Without massive trauma of the carotid bifurcation, histological alterations cannot be expected. Without signs of rapid death, findings of acute circulatory failure, macromorphological and micromorphological findings of neck compression, and reliable markers indicating relevant impact on the carotid bifurcation the diagnosis of a lethal reflex cannot be verified.Key pointsAmong 22 cases of strangulation causing death, there were 16 cases of hanging and 6 cases of ligature strangulationFew cases showed small haemorrhages located predominantly in the surrounding fat and connective tissues; however, the haemorrhages did not have any effects on the tissuesNeck compression had minimal effects on heat shock protein 27 expression in carotid artery tissueAquaporin-3 staining suggested it is not a useful marker for relevant neck pressure, or that there had not been any relevant neck impactOur findings suggested no direct evidence for reflex cardiac death resulting from a brief force against the neck.

Evaluation of RapidSTAT®, DrugWipe® 6S, DrugScreen® 5TK and DrugScreen® 7TR for on-site drug testing in German police roadside traffic patrol.

Driving under the influence of drugs (DUID) remains a subject of concern worldwide, and its increasing trend is likely to continue. Therefore, there is a constant need for reliable on-site drug tests to identify drugged drivers during roadside patrols. Performance and reliability of four on-site drug tests were evaluated among a high number of DUID cases in Germany. Results of oral fluid (OF) (RapidSTAT® and DrugWipe® 6S) and urine (DrugScreen® 5TK and 7TR) test devices were compared with corresponding serum/plasma results obtained by confirmation analyses in consideration of recommended analytical limits for substances pertaining the annex of the German Road Traffic Code ('Straßenverkehrsgesetz', StVG) s. 24a (2). Overall, the screening devices performed well for individual drugs; however, none of the test devices assessed in this study fulfilled the ROSITA-1 criteria (sensitivity, specificity ≥ 90% and accuracy ≥ 95%) for all substances. Our data demonstrated that both urine tests showed high sensitivities for most compounds. DrugWipe® 6S (94%) and RapidSTAT® (93%) revealed high sensitivities, especially for amphetamine screening. Poor specificities (<90%) and accuracies (<95%) were observed for all tests except for low-prevalent substances (e.g., opiates). For drug testing in OF, Δ9 -tetrahydrocannabinol (THC) still seems to be a compound of concern due to poor sensitivity (RapidSTAT®, 77%; DrugWipe® 6S, 85%), although the results indicate improvements compared with previously reported data. Although the obtained data indicate reliable detection for some substances, deployment of trained police officers is inevitable to identify DUID suspects by signs of recent use and recognising impairment.

Fatty acid esters as novel metabolites of γ-hydroxybutyric acid: A preliminary investigation.

γ-Hydroxybutyric acid (GHB) is a substance frequently abused as a knockout agent. Because of possible amnesia experienced by victims of GHB exposure and the short detection time of GHB in biological samples, the proof of GHB uptake is often challenging for forensic toxicologists. For this reason, various approaches have been evaluated to prolong the detection of GHB intake. In the present study, a fatty acid ester of GHB (4-palmitoyloxy butyrate [GHB-Pal; 3-carboxypropyl hexadecanoate]) was synthesized with the intent of examining whether such esters could be detected as metabolites of GHB in blood samples. Using the structurally elucidated synthesis product (structural elucidation by means of high performance liquid chromatography quadrupole time of flight mass spectrometry [LC-QToF-MS]), an LC triple quadrupole mass spectrometric (LC-MS/MS) method was established for the detection of GHB-Pal. Blood (plasma) samples from four cases in which GHB was previously detected at relevant concentrations (56.1-96.5 µg/ml) were analyzed with respect to GHB-Pal. Signals for GHB-Pal, as well as possible signals for other fatty acid esters of GHB, were detectable in these specimens. (Negative) control samples (20 plasma samples and 20 red blood cell/blood clot samples; from cases in which an intake of GHB or its precursors was not assumed) were all negative for GHB-Pal. To evaluate a possible forensic benefit of GHB fatty acid esters (prolongation of the detection window of a GHB uptake), the analysis of additional plasma samples collected after GHB uptake (or controlled GHB administration) and quantification of GHB fatty acid esters are needed.

Case report: fatal bleeding from a duodenal ulcer-Dieulafoy's lesion?

A 46-year-old man was admitted to the hospital by ambulance due to syncope. A standard blood screening showed a normal Hb value. The man had known hemorrhoids and a single fresh rectal bleeding earlier at home. On the following morning, the patient suddenly required resuscitation within a few minutes and subsequently died. Autopsy revealed a fatal hemorrhage with blood loss in the stomach and small and large intestines and a mucosal defect of the duodenum. After autopsy, the question arose whether the cause of death might have been a rare Dieulafoy's lesion-aim of this case report was to clarify the diagnosis.

Comparative Study: Postmortem Long-Term Stability of Endogenous GHB in Cardiac Blood, Femoral Blood, Vitreous Humor, Cerebrospinal Fluid and Urine with and without Sodium Fluoride Stabilization.

The interpretation of postmortem γ-hydroxybutyric acid (GHB) concentrations is challenging due to endogenous existence and postmortem GHB production in body tissues and fluids. As an additional complication, formation of GHB was also described in stored postmortem samples. We examined cardiac blood, femoral blood, vitreous humor, cerebrospinal fluid and urine of eight different corpses (male/female 5/3, aged 33-92 years, postmortem interval 1-6 days) where no intake of GHB or one of its precursors was assumed. All samples were collected during autopsy and divided into two aliquots. To one of the aliquots, sodium fluoride (NaF, 1% w/v) was added. Both aliquots were vortexed, further divided into seven aliquots and stored at -20°C. GHB concentrations were measured immediately and subsequently 1 day, 7 days, 2 weeks, 4 weeks, 3 months and 6 months, after sample collection using trimethylsilyl derivatization and gas chromatography, coupled to single quadrupole mass spectrometry. Similar progression curves of GHB concentrations were obtained for the different matrices in the individual corpses. Femoral and cardiac blood GHB concentrations were always found to be higher than in vitreous humor, cerebrospinal fluid, and urine irrespective of stabilization and storage time. None of the obtained GHB concentrations exceed the cutoff values for postmortem matrices commonly used for the identification of an exogenous GHB intake (urine, venous blood and cerebrospinal fluid: 30 mg/L, cardiac blood and vitreous humor 50 mg/L). No significant differences were found for the GHB concentrations measured immediately and 6 months after autopsy. However, we found a significant increase for the GHB concentrations 4 weeks as well as 3 months after sample collection, which was followed by a decrease nearly to initial values. There were no significant differences between samples with and without NaF addition. The data presented are useful for the interpretation of GHB concentrations in upcoming death cases, with special attention to storage conditions and different postmortem matrices.

Expression of heat shock proteins 27, 60, and 70 in amphetamine and cocaine associated deaths.

INTRODUCTION: The hearts of amphetamine and cocaine users demonstrate essentially the same microscopic features: hypertrophy, interstitial fibrosis, myocyte hypertrophy and intimal and medial hyperplasia. According to Karch (2016), some investigations suggest that amphetamines have properties that make users less likely to experience myocardial infarction than cocaine users. The exposure to amphetamine is associated with the production of heat shock proteins (HSP) whereas cocaine is not. Not all the HSP are present in normal living conditions of cells but their expression is increased when cells are exposed to stress, like heat, anoxemia, and ischemia. It has been known before that increased HSP production is a myocardial response in adaptation to cardiac ischemia and that the production of HSP might influence myocardial resistance to infarction. Furthermore, production of HSP is an explanation of the known ability of amphetamines to cause hyperthermia. The hypothesis of a cytoprotective function of HSP in amphetamine-associated deaths in comparison to cocaine-associated deaths and controls was investigated. MATERIAL AND METHODS: Study group: 39 amphetamine-related fatal cases, 27 cocaine-associated deaths. CONTROL GROUP: 42 cases with other causes of death. Immunohistochemical staining of HSP 27, HSP 60, and HSP 70 in heart, liver, and kidney. RESULTS: 16 out of 39 (41.0%) amphetamine-related fatal cases showed a positive HSP expression, predominantly HSP 70 in myocardial tissue. In cocaine-associated deaths 15 out of 27 (55.5%) cases were positive, also mainly HSP 70. In the kidney in amphetamine-associated deaths 18 out of 39 (46.1%) cases were positive, in cocaine-associated deaths 21 out of 27 (77.7%) cases. The cocaine group showed significantly increased expression for HSP 27 and 70 in the liver and HSP 70 in the kidney compared to the control as well as amphetamine group. Furthermore, the cocaine group showed significantly increased expression for HSP 27 and 70 in the heart compared to the control but not the amphetamine group. CONCLUSION: The hypothesis of Karch that in amphetamine-associated deaths a positive HSP expression has in contrast to cocaine-related deaths a cytoprotective function cannot be verified. Furthermore, cocaine and benzoylecgonine seem to independently lead to an increased expression of HSP 27 both in the liver and in the heart.

Efficacy of Δ9 -Tetrahydrocannabinol (THC) Alone or in Combination With a 1:1 Ratio of Cannabidiol (CBD) in Reversing the Spatial Learning Deficits in Old Mice.

Decline in cognitive performance, an aspect of the normal aging process, is influenced by the endocannabinoid system (ECS). Cannabinoid receptor 1 (CB1) signaling diminishes with advancing age in specific brain regions that regulate learning and memory and abolishing CB1 receptor signaling accelerates cognitive aging in mice. We recently demonstrated that prolonged exposure to low dose (3 mg/kg/day) Δ9-tetrahydrocannabinol (THC) improved the cognitive performances in old mice on par with young untreated mice. Here we investigated the potential influence of cannabidiol (CBD) on this THC effect, because preclinical and clinical studies indicate that the combination of THC and CBD often exhibits an enhanced therapeutic effect compared to THC alone. We first tested the effectiveness of a lower dose (1 mg/kg/day) THC, and then the efficacy of the combination of THC and CBD in 1:1 ratio, same as in the clinically approved medicine Sativex®. Our findings reveal that a 1 mg/kg/day THC dose still effectively improved spatial learning in aged mice. However, a 1:1 combination of THC and CBD failed to do so. The presence of CBD induced temporal changes in THC metabolism ensuing in a transient elevation of blood THC levels. However, as CBD metabolizes, the inhibitory effect on THC metabolism was alleviated, causing a rapid clearance of THC. Thus, the beneficial effects of THC seemed to wane off more swiftly in the presence of CBD, due to these metabolic effects. The findings indicate that THC-treatment alone is more efficient to improve spatial learning in aged mice than the 1:1 combination of THC and CBD.

Follow up: palmitic acid ester of tetrahydrocannabinol (THC) and palmitic acid diester of 11-hydroxy-THC - unsuccessful search for additional THC metabolites.

OBJECTIVES: In a previous investigation we searched for the occurrence of palmitic acid ester compounds of delta9-tetrahydrocannabinol (THC) and its primary metabolite 11-hydroxy-delta9-THC (11-OH-THC) in human body fluids and tissues (THC palmitic acid monoester [THC-Pal] and 11-OH-THC palmitic acid diester [11-OH-THC-DiPal]). As those esters could not be detected in various human body fluids (e.g. blood) or tissues (e.g. adipose tissue) we extended the investigation analyzing adipose tissue samples of mice previously given synthetic THC or a cannabis extract. METHODS: In total, 48 adipose tissue samples previously tested positive for THC by means of a liquid chromatographic triple quadrupole mass spectrometric (LC-QQQ-MS) method were analyzed for the presence of THC-Pal and 11-OH-THC-DiPal by means of LC-QQQ-MS. RESULTS: THC-Pal and 11-OH-THC-DiPal were not detected among the adipose tissue samples analyzed despite the presence of high THC concentrations within the adipose tissue. THC concentrations in adipose tissue were in a range of approximately 7-2,595 ng/g (median: 468 ng/g, average: 704 ng/g). CONCLUSIONS: A (site-specific) synthesis of 11-OH-THC palmitic acid monoesters (11-hydroxy-delta9-THC-1-palmitate and 11-palmitoyloxy-delta9-THC) still remains to be done. After synthesis of these monoesters, their presence in the body fluids and tissues after THC administration should be investigated.

Factitious disorders in Germany-a detailed insight.

Factitious disorders (FDs) are well known to a majority of physicians; however, the corresponding ICD-10 diagnosis F68.1 remains severely under assigned and often misdiagnosed. Based on a previously conducted nationwide survey in Germany, we extended the analyzed variables to further understand FD characteristics.The assignments regarding the following variables in the German diagnosis-related group statistics were analyzed: residence of the patient and location of the diagnosing institution, primary referral to the diagnosing institution, reason for admission and discharge, specialty department, total length of stay, length of stay in the longest treating department, surgery performed, case mix revenue, regional type of the treating institution, and patients' region of origin.A very distinct difference was observed in the assignment rates based on the homeland of the diagnosed patient and diagnosing institution. The assignment rate showed no significant difference across German regions. Based on our findings, a patient with FD in Germany might exhibit the following "typical" traits: A woman in her late thirties from a rural area is referred by a physician or another hospital wherein she was previously treated for more than a day to an institution for fully inpatient hospital treatment wherein she completes her treatment regularly. Dermatology, neurology, emergency, and internal medicine departments tend to be confronted with patients with FDs more often than other departments; however, surgery is performed in every fifth case. Patients are primarily treated in only one department for ~ 25 days. The case mix revenue will most probably not exceed €5000.

Fox sign in a case of terminal stage pancreatic cancer and suggestions for diagnosis.

Skin signs in acute pancreatitis are well-known and frequently discussed manifestations accompanied by unfavorable prognoses although they may rarely appear in clinical and forensic medicine. In 2018, the district attorney's office ordered a forensic autopsy for a 74-year-old man with terminal stage pancreatic cancer. The autopsy was ordered based on accusations of the deceased's widow regarding alleged medical malpractice and poor hospital care. The widow filed a grievance about multiple unsuccessful attempts to draw blood from her husband in addition to a diaper dermatitis at the right groin. An autopsy and additional histological examinations were performed. After considering all findings, the diaper dermatitis was eventually assumed to be a Fox sign caused by acute pancreatitis, and the allegations of medical malpractice were refuted. This case led us to identify another case with suspected cutaneous manifestations in pancreatic disease. We performed immunohistochemical staining on those two cases and six control cases to examine whether there was detectable presence of pancreatic lipase and trypsin in the skin discolorations and whether it could be used as a feasible method to verify skin signs associated with pancreatitis. Based on our findings, a minor disseminated lipase and trypsin staining should be considered regular and is therefore not conclusive of a skin sign associated with pancreatitis. Moreover, trypsin does not seem to be as suitable as lipase for this suggested immunohistochemical method. Nevertheless, this method might be a useful addition for determining the origin of skin discoloration and verifying skin signs associated with pancreatitis.