Seminal plasma (SP) induces a rapid transforming growth factor beta 1 (TGFß1)-independent up-regulation of epithelial-mesenchymal transdifferentiation (EMT) and myofibroblastic metaplasia-markers in endometriotic (EM) and endometrial cells.

PURPOSE: To study if short-term exposure (2 h and 6 h) of endometrial/endometriotic tissues and cells to 10% seminal plasma (SP) can induce EMT/metaplasia. METHODS: Basic research experimental study was carried out in a University hospital-based fertility center. Semen samples, peritoneal fluid (PF) from endometriosis patients, endometrial biopsy from premenopausal women, immortalized endometriotic epithelial cell line (12Z), and immortalized endometrial stromal cell line (St-T1b) were studied. Rapid stain identification test (RSID), TGFß1 immunofluorescence of washed sperms, TGFß1-ELISA of SP and PF, in vitro study (2 h and 6 h incubation) and real-time PCR of endometrial tissue and cell lines to analyze gene expression of EMT/metaplasia markers and mediators were done. RESULTS: SP is still detectable in washed semen. TGFß1 was expressed on the plasma membrane of the sperms and was significantly more concentrated in SP (88.17 ng/ml) than PF. 10% SP induced an up-regulation of alpha smooth muscle actin expression in endometrial tissue (p = 0.008) and in 12Z cells (p = 0.05), mostly TGFß1-independent. TWIST expression was persistently significantly down-regulated while Snail1 and 2 were up-regulated, though insignificant. CONCLUSION: Our results provide novel evidence to support that even in semen washed twice, SP is still detectable. The changes in EMT/metaplasia markers and mediators give a new insight into a possible effect of SP on the pathogenesis of endometriosis.

RNA/DNA co-analysis from human skin and contact traces--results of a sixth collaborative EDNAP exercise.

The European DNA profiling group (EDNAP) organized a sixth collaborative exercise on RNA/DNA co-analysis for body fluid/tissue identification and STR profiling. The task was to identify skin samples/contact traces using specific RNA biomarkers and test three housekeeping genes for their suitability as reference genes. Eight stains, a skin RNA dilution series and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 22 participating laboratories using RNA extraction or RNA/DNA co-extraction methods. Two sets of previously described skin-specific markers were used: skin1 pentaplex (LCE1C, LCE1D, LCE2D, IL1F7 and CCL27) and skin2 triplex (LOR, KRT9 and CDSN) in conjunction with a housekeeping gene, HKG, triplex (B2M, UBC and UCE). The laboratories used different chemistries and instrumentation. All laboratories were able to successfully isolate and detect mRNA in contact traces (e.g., human skin, palm-, hand- and fingerprints, clothing, car interiors, computer accessories and electronic devices). The simultaneous extraction of RNA and DNA provides an opportunity for positive identification of the tissue source of origin by mRNA profiling as well as a simultaneous identification of the body fluid donor by STR profiling. The skin markers LCE1C and LOR and the housekeeping gene marker B2M were detected in the majority of contact traces. Detection of the other markers was inconsistent, possibly due to the low amounts and/or poor quality of the genetic material present in shed skin cells. The results of this and the previous collaborative RNA exercises support RNA profiling as a reliable body fluid/tissue identification method that can easily be combined with current STR typing technology.

Mitochondrial deoxyribonucleic acid may play a role in a subset of sudden infant death syndrome cases.

AIM: It has been suggested that progressive adenosine triphosphate (ATP) depletion could play a key role in sudden infant death syndrome (SIDS). Because mitochondrial deoxyribonucleic acid (mtDNA) codes for a subset of essential genes for oxidative phosphorylation, we investigated 22 mtDNA polymorphisms in a large sample of Caucasian SIDS cases. METHODS: A total of 774 samples were analysed, 365 from infant SIDS cases (mean age 131 days) and 409 from controls. These were investigated for the presence of 22 haplogroup-specific single nucleotide polymorphisms (SNPs), using a SNaPshot assay, a mini-sequencing assay that combines polymerase chain reaction (PCR) and sequencing. RESULTS: No significant differences in assigned haplogroups could be detected between the groups. With regard to gender and age, we found significant correlations for SNP positions 3010, 8251, 13 708, 14 470, 15 904 and 16 519. The most prominent result was the A allele in SNP 14 470 in male SIDS cases (p = 0.01). CONCLUSION: This is the largest study on mtDNA polymorphisms in SIDS to date, and our results indicate that mtDNA may play a role in a subset of SIDS cases. In order to complement these significant results, it is important to consider nuclear gene coding for mitochondrial proteins in future studies.

Evidence based palaeopathology: meta-analysis of Pubmed-listed scientific studies on pre-Columbian, South American mummies.

The aim of this study was to review all Pubmed(®)-listed palaeopathological studies performed on pre-Columbian South American mummies. A total of 61 studies were found (1977-2005). Review criteria included e.g. method of examination, method of mummification, palaeopathological diagnoses and individual age of mummies as well as dating, which ranged from 7500 to 500 years BP, mainly (if reported) originating from the Chiribaya and Chinchorro cultures. The average age of the 99 individually reported mummies was about 25 years. Only six studies included computed tomography, thirteen studies used classical radiography as an examination method. Three studies analysed parasite related diseases, especially caused by Trypanosoma cruzi. Among all of the reported infectious diseases (n=9), there were seven studies presenting cases of tuberculosis. The results were also compared inter-culturally. In 61 studies (37 epidemiological and 24 case reports) more than 6400 mummified individuals were analysed. By contrast, meta-analytic data for ancient Egyptian mummies (Zweifel et al., 2009) included about 3000 analysed individuals in 131 studies (85 case reports and 46 epidemiological studies). In general, ancient Egyptian mummies were shown to be intentionally mummified, whereas the Pre-Columbian American mummies showed a great diversity of spontaneous mummification. However, ritualistic mummification methods were also practised (n=2). This study's results shall assist to improve evidence-based research in palaeopathology.

RNA/DNA co-analysis from human menstrual blood and vaginal secretion stains: results of a fourth and fifth collaborative EDNAP exercise.

The European DNA Profiling Group (EDNAP) organized a fourth and fifth collaborative exercise on RNA/DNA co-analysis for body fluid identification and STR profiling. The task was to identify dried menstrual blood and vaginal secretion stains using specific RNA biomarkers, and additionally test 3 housekeeping genes for their suitability as reference genes. Six menstrual blood and six vaginal secretion stains, two dilution series (1/4-1/64 pieces of a menstrual blood/vaginal swab) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 24 participating laboratories, using RNA extraction or RNA/DNA co-extraction methods. Two novel menstrual blood mRNA multiplexes were used: MMP triplex (MMP7, MMP10, MMP11) and MB triplex (MSX1, LEFTY2, SFRP4) in conjunction with a housekeeping gene triplex (B2M, UBC, UCE). Two novel mRNA multiplexes and a HBD1 singleplex were used for the identification of vaginal secretion: Vag triplex (MYOZ1, CYP2B7P1 and MUC4) and a Lactobacillus-specific Lacto triplex (Ljen, Lcris, Lgas). The laboratories used different chemistries and instrumentation and all were able to successfully isolate and detect mRNA in dried stains. The simultaneous extraction of RNA and DNA allowed for positive identification of the tissue/fluid source of origin by mRNA profiling as well as a simultaneous identification of the body fluid donor by STR profiling, also from old and compromised casework samples. The results of this and the previous collaborative RNA exercises support RNA profiling as a reliable body fluid identification method that can easily be combined with current STR typing technology.

Accidental mechanical asphyxia of children in Germany between 2000 and 2008.

Accidents constitute one of the greatest risks to children, yet there are few medical reports that discuss the subject of accidental asphyxia. However, a systematic analysis of all documented cases in Germany over the years 2000-2008 has now been conducted, aiming at identifying patterns of accidental asphyxia, deducing findings, defining avoidance measures and recommending ways of increasing product safety and taking possible precautions. The analysis is based on a detailed retrospective analysis of all 91 relevant autopsy reports from 24 different German forensic institutes. A variety of demographic and morphological data was systematically collected and analysed. In 84 of the 91 cases, the sex of the victim was reported, resulting in a total of 57 boys (68 %) and 27 girls (32 %). The age spread ranged between 1 day and 14 years, with an average of 5.9 years. Most accidents occurred in the first year of life (20 %) or between the ages of 1 and 2 years (13 %). In 46 % of cases, the cause of death was strangulation, with the majority occurring in the home environment. In 31 % of all cases, the cause of death was positional asphyxia, the majority resulting from chest compression. In 23 % of cases, the cause of death was aspiration, mainly of foreign bodies. Today, accidental asphyxiation is a rare cause of death in children in Germany. Nevertheless, the majority of cases could have been avoided. Future incidence can be reduced by implementing two major precautions: increasing product safety and educating parents of potentially fatal risks. Specific recommendations relate to children's beds, toys and food.

Prevalence of chronic migraine and medication overuse headache in Germany--the German DMKG headache study.

Population-based epidemiological studies about the prevalence of chronic migraine using the 2004 International Headache Society (IHS) classification definition are rare. We analysed the data of the Deutsche Migräne und Kopfschmerz Gesellschaft headache study, which included 7417 adults in three regions of Germany, with respect to their headache. Additionally, body mass index, alcohol consumption and smoking behaviour were recorded. Using the IHS definition from 2004, chronic migraine was diagnosed in 0.2% of the population. Half of these patients also fulfilled the criteria of medication overuse headache (MOH). The distribution of migraine attacks per subject was highly skewed, with only 14% of all migraine patients having more than six migraine attacks per month. Patients with chronic migraine or MOH seem more often to be active smokers than controls without headache. A body mass index of ≥ 30 was present significantly more often in patients with MOH than in controls or in patients with episodic migraine. The skewed distribution of the numbers of attacks per patient supports the recommendation to differentiate between episodic migraine with low and high attack frequency, as is done in the classification of tension-type headache. It further suggests that migraine with high attack frequency might be biologically different. The higher prevalence of smokers and of patients with a body mass index ≥ 30 in chronic migraine or MOH supports the idea of a frontal dysfunction in these patients.

Does breastfeeding reduce the risk of sudden infant death syndrome?

BACKGROUND: In the last 20 years, the prevention campaigns to reduce the risk of sudden infant death syndrome were very successful. In some countries the advice to breastfeed is included in the campaigns' messages, but in other countries it is not. OBJECTIVE: To examine the association between type of infant feeding and sudden infant death syndrome. METHODS: The German Study of Sudden Infant Death is a case-control study of 333 infants who died of sudden infant death syndrome and 998 age-matched controls. RESULTS: A total of 49.6% of cases and 82.9% of controls were breastfed at 2 weeks of age. Exclusive breastfeeding at 1 month of age halved the risk, partial breastfeeding at the age of 1 month also reduced the risk of sudden infant death syndrome, but after adjustment this risk was not significant. Being exclusively breastfed in the last month of life/before the interview reduced the risk, as did being partially breastfed. Breastfeeding survival curves showed that both partial breastfeeding and exclusive breastfeeding were associated with a reduced risk of sudden infant death syndrome. CONCLUSIONS: This study shows that breastfeeding reduced the risk of sudden infant death syndrome by approximately 50% at all ages throughout infancy. We recommend including the advice to breastfeed through 6 months of age in sudden infant death syndrome risk-reduction messages.

Regional variations in the prevalence of migraine and tension-type headache applying the new IHS criteria: the German DMKG Headache Study.

The prevalence of migraine and tension-type headache (TTH) varies considerably with respect to gender, age group and geographic regions. Methodological differences in the assessment and classification of cases are a major cause of this variability across studies, limiting the ability to perform true regional comparisons. We conducted three population-based studies in different German regions and assessed headache prevalence and headache characteristics in face-to-face interviews, applying standardized methods. We analysed the 6-month prevalence of migraine, TTH and their probable subtypes based on the new criteria of the International Headache Society (IHS). Among the 7417 participants in all three regions, the pooled 6-month prevalence of migraine, probable migraine, TTH and probable TTH was 6.75, 4.40, 19.86 and 11.61%, respectively. Despite the application of standardized classification methods, regional variations between 4.39 and 8.00% for migraine and 15.44 and 23.64% for TTH were observed, indicating differences in the local headache burden. Application of the new IHS criteria yielded headache categories that were not mutually exclusive, indicating a need for further discussion about the value of probable headache types in epidemiological studies.

Prone sleeping position increases the risk of SIDS in the day more than at night.

BACKGROUND: SIDS mortality is higher during the night than in the day. AIM: (1) To examine risk factors for SIDS by time of day and (2) to see if the proportion of deaths at night has changed from prior to the 'Back to Sleep' campaign, which recommended infants sleep supine. METHODS: A large population-based SIDS matched case-control (GeSID) study conducted from 1998 to 2001 (when the prevalence of infants placed prone to sleep was 4.1%). The reference sleep of the controls was matched for the estimated time of death for the case. Risk factors for SIDS were examined for night-time and day-time deaths. The estimated time of death was compared with that from an earlier study in Germany (1990-1994 when prevalence of prone sleeping was 32.2%). RESULTS: There were 333 SIDS cases and 998 matched controls. The increased risk with placed prone to sleep was significantly different during the day [adjusted OR = 18.15 (95% CI = 5.91-55.69)] compared with during the night [adjusted OR = 3.49 (95% CI = 1.46-8.39; p-value for interaction = 0.011)]. There was no significant difference in the other risk factors examined by time of day in the multivariate analysis. The mean time found dead was 09:07. In the earlier study the mean time found dead was 08:54 and the difference was not significant (p = 0.57). CONCLUSIONS: This study confirms previous observations that prone sleeping position carries a greater risk during the day than at night. However, the reduction in infants sleeping prone has not been associated with a reduced number of deaths in the day in Germany.

Nicotine and cotinine in infants dying from sudden infant death syndrome.

The aim of this component of the German Study on Sudden Infant Death was to determine (1) nicotine concentrations in hair (NCH), as a marker of long standing exposure to tobacco, (2) cotinine concentrations in pericardial fluid (CCP) and (3) cotinine concentrations in liquor cerebrospinalis (CCL), the latter measures being markers of recent exposure to tobacco in the last few hours of life. The results obtained were compared with data on parental smoking revealed from interviews. In 100 cases of sudden infant death syndrome, material was taken at autopsy to determine NCH. In 41 cases, NCH and CCP, and in 70 cases, NCH and CCL were determined. Infants of mothers who stated having smoked during pregnancy had higher NCH than infants of non-smoking mothers (p = 0.008). Furthermore, there was a weak but statistically significant relationship between NCH's and the daily cigarette consumption of the mother during pregnancy (n = 64, r = 0.24, p = 0.05). In 43% of infants, nicotine could be detected in their hair, although the mothers had said at the interview that they did not smoke during pregnancy. On the other hand, in 33% of infants whose mother stated they had smoked during pregnancy nicotine was not detectable in the infant's hair. CCP's were strongly correlated with CCL's (r = 0.62, p = 0.0027). For this reason, both parameters were treated as equivalent for the detection of tobacco smoke exposure in the last hours before death. The influence of breast-feeding was evaluated by comparison of the nicotine concentrations in breast fed and non-breast-fed infants from smokers and non-smokers. Fivefold higher nicotine concentrations were determined in non-breast-fed infants of parents who smoked as compared to all other groups. It can be concluded that nicotine intake by passive smoking is much more important than by breast-feeding. We conclude that both interview data and biochemical measures should be sought to understand the true exposure to tobacco smoke.

Do immunisations reduce the risk for SIDS? A meta-analysis.

BACKGROUND: There are claims that immunisations cause sudden infant death syndrome (SIDS), but some studies have found either no association or that they are associated with a reduced risk of SIDS. AIMS: To conduct a meta-analysis examining the relationship between immunisation and SIDS. METHODS: Nine case-controls studies were identified examining this association, of which four adjusted for potential confounders. RESULTS: The summary odds ratio (OR) in the univariate analysis suggested that immunisations were protective, but the presence of heterogeneity makes it difficult to combine these studies. The summary OR for the studies reporting multivariate ORs was 0.54 (95% CI=0.39-0.76) with no evidence of heterogeneity. CONCLUSIONS: Immunisations are associated with a halving of the risk of SIDS. There are biological reasons why this association may be causal, but other factors, such as the healthy vaccinee effect, may be important. Immunisations should be part of the SIDS prevention campaigns.

Headache prevalence among adolescents--the German DMKG headache study.

This population-based cross-sectional study examined the 3-month prevalence of headache, migraine and tension-type headache (TTH) among adolescents aged 12-15 years in Germany. Students (n = 3324) from 20 schools completed a questionnaire on general and headache-specific pain which included a sociodemographic module. The headache-specific questionnaire complied with the respective revised criteria of the International Headache Society (IHS). 'Modified criteria' changed the item 'duration' in migraine (>30 min instead of > 4 h). The overall 3-month prevalence of headache was 69.4% (boys 59.5%, girls 78.9%), with 4.4% of the adolescents suffering from frequent (>or=14 days/3 months) and severe (grade 8-10 on a 10-point visual analogue scale) headache and 1.4% (boys 0.9%, girls 1.9%) from headache >or= 15 days/month. The 3-month prevalence of migraine was 2.6% (boys 1.6%, girls 3.5%) applying strict IHS criteria and 6.9% (boys 4.4%, girls 9.3%) with modified criteria; 12.6% (boys 8.3%, girls 16.7%) suffered from probable migraine, 0.07% fulfilled the criteria for chronic migraine, 4.5% (boys 4.6%, girls 4.3%) suffered from TTH, 0.2% from chronic TTH and 15.7% (boys 14.5%, girls 16.9%) from probable TTH. Headache and migraine were more common in girls than in boys and in teenagers, especially in girls, aiming at higher education. Recurrent headache and primary headache disorders are common complaints among German adolescents, especially among girls.

Sudden infant death syndrome: no increased risk after immunisation.

BACKGROUND: Although previous studies have shown either no association between immunisation and SIDS or even a decreased risk of SIDS, adverse effects, including death, from immunisations continue to cause concern, especially when a new vaccine is introduced. METHODS: A large case control study with immunisation data on 307 SIDS cases and 971 controls. RESULTS: SIDS cases were immunised less frequently and later than controls. Furthermore there was no increased risk of SIDS in the 14 days following immunisation. There was no evidence to suggest the recently introduced hexavalent vaccines were associated with an increased risk of SIDS. CONCLUSIONS: This study provides further support that immunisations may reduce the risk of SIDS.

Do risk factors differ between explained sudden unexpected death in infancy and sudden infant death syndrome?

BACKGROUND: In Germany, 2910 infants died in 2004; for many infants the reason was clear, especially prematurity or congenital abnormalities. However, 394 babies die every year suddenly and unexpectedly. The cause may be immediately clear, but is often not obvious. AIMS: (1) To describe the causes of explained sudden unexpected death in infancy (SUDI) and (2) to compare risk factors for sudden infant death syndrome (SIDS) and explained SUDI. METHODS: A 3-year population-based case-control study in Germany, 1998-2001. RESULTS: 455 deaths, of which 51 (11.2%) were explained. Most of these deaths were due to respiratory or generalised infections. The risk factors for SIDS and explained SUDI were remarkably similar except for sleep position and breast feeding. Prone sleeping position is a major risk factor for SIDS (adjusted odds ratio (OR) 7.16, 95% confidence interval (CI) 3.85 to 13.31) but not for explained SUDI (adjusted OR 1.71, 95% CI 0.25 to 11.57). Not being breast fed in the first 2 weeks of life is a risk factor for SIDS (adjusted OR 2.37, 95% CI 1.46 to 3.84) but not for explained SUDI (adjusted OR 0.39, 95% CI 0.08 to 1.83). CONCLUSIONS: Prone sleeping position is a unique risk factor for SIDS. Socioeconomic disadvantage and maternal smoking are risk factors for both SIDS and explained SUDI, and provide an opportunity for targeted intervention.

Are autopsies of help to the parents of SIDS victims? A follow-up on SIDS families.

Little is known about what bereaved parents feel about the autopsy performed on their child. A multi-centre case control study of sudden infant death syndrome (SIDS) victims was carried out in Germany between 1998 and 2001, in which all infants had been autopsied. We performed a follow-up study 4-7 years after the parents had lost their child. A total of 141 parents filled in the questionnaire, which were sent to them by the study centre. Of these, 71% had had another child after the SIDS/sudden unexpected death in infancy. The majority (83%) of the participating parents found the autopsy helped them to cope better with the death. A large proportion (46%) did not want any professional help after the death, and 55% did not wish to have any contact with a self-help group. We conclude that the autopsy is helpful to the majority of bereaved parents. Professional help and self-help groups should be offered to the parents even if the majority in our study did not want to use either.

Prevention of sudden infant death syndrome (SIDS) due to an active health monitoring system 20 years prior to the public "Back to Sleep" campaigns.

BACKGROUND: Before reunification, the post-neonatal mortality rate was lower in East Germany than in West Germany. Moreover, the incidence of SIDS (sudden infant death syndrome) was much lower in the East. METHODS: Mortality data on sudden infant death syndrome (SIDS) from West and East Germany since 1980 as well as post-neonatal mortality data for both states since 1970 were examined. 95% Confidence intervals were calculated for the rates. Witnesses from the former East Germany who were involved at the time were also interviewed and archives were searched. RESULTS: We found that as early as 1972 active monitoring of infant and child mortality rates in East Germany had shown that the prone sleeping position was dangerous for infants: the post-neonatal mortality rate was approximately 1 per 1000 live births lower in East than in West Germany during the 20 years before reunification. In contrast, in the West, prone sleeping was only discovered to be a risk factor for SIDS in the early 1990s. CONCLUSIONS: Active monitoring is an effective tool in the early detection of risk factors and serves to prevent unnecessary deaths.

The San Diego definition of SIDS: practical application and comparison with the GeSID classification.

The new definition of the term sudden infant death syndrome (SIDS) and the criteria introduced in San Diego for the subclassification of cases have been used to re-classify the first 100 consecutive cases of sudden and unexpected infant deaths that were registered with the German SIDS study (GeSID). Although there are 30 different variables that have to be considered in the general and stratified sections of the San Diego definition, it is practical, in particular, as an international standard to perform scientific studies. The comparison of the San Diego definition and the classification used for GeSID shows similarities in the methods but differences in the criteria used. Nevertheless, the numbers of cases classified as SIDS and borderline SIDS are similar (San Diego n=69, GeSID n=74). The SIDS IA criteria of the San Diego definition were not fulfilled by any case because metabolic screening and vitreous chemistry were not included in the GeSID investigation scheme. An important advantage of the San Diego definition is the introduction of the category of unclassified sudden infant death, which includes cases for which no autopsy was performed. This demonstrates that such cases cannot be classified as SIDS. In conclusion, we recommend the universal acceptance and use of the San Diego SIDS definition.

Infection, health problems, and health care utilisation, and the risk of sudden infant death syndrome.

AIM: To examine whether symptoms suggestive of infection, health problems, and health care utilisation are risk factors for SIDS. METHODS: Matched case-control study with 333 SIDS infants and 998 control infants matched for region, age, gender, and reference sleep. Information was obtained by parental interview, paediatrician completed questionnaire, and hospital admission data. RESULTS: No symptoms were associated with SIDS after adjustment for potential confounders. Illness in the last four weeks as reported by the paediatrician did not differ between cases and controls. Developmental problems and special investigations at any stage of life significantly increased the risk of SIDS (adjusted OR = 2.14 and 2.07). Admission to hospital after the first week of life was associated with an increased risk of SIDS (adjusted OR = 1.88). CONCLUSION: Symptoms of infection and illness are no longer risk factors for SIDS in communities such as Germany where few infants sleep prone. The increased risk of SIDS with developmental problems may indicate that infants which subsequently die of SIDS are abnormal or in some way vulnerable.

Unnatural causes of sudden unexpected deaths initially thought to be sudden infant death syndrome.

The aim of this clinicopathological study was to determine the frequency of infant deaths due to unnatural causes among cases of sudden and unexpected infant death. Nine institutes of legal medicine in Germany that took part in the German study on Sudden Infant Death Syndrome (GeSID), representing 35% of the German territory, investigated in a 3-year period (from 1998 to 2001) 339 cases of infant death that were not expected to be due to unnatural causes from the first external examination. All cases were investigated by complete, standardised, post-mortem examination including death scene investigation, autopsy, histology, toxicology and neuropathology. The frequency of unnatural deaths was 5.0% (n=17). The causes of death were head injury (n=7), suffocation (n=5), poisoning (n=2), neglect (n=2) and septicaemia due to aspiration of a foreign body (n=1). Two deaths were unsuspected accidents and 12 were due to infanticide. In 3 cases, it was not possible to differentiate between accidental death and infanticide. A complete postmortem examination including an analysis of the clinical history, death scene investigation, autopsy, histology, toxicology, and neuropathology is mandatory to differentiate sudden and unexpected deaths due to natural causes (e.g. SIDS) and cases of unnatural death.