Putrefaction: An Additional Complicating Factor in the Assessment of Freshwater Drownings in Rivers

Background hypothesis: That bodies immersed in freshwater (rivers) may show greater changes of decomposition than bodies immersed in saltwater (sea). Fifty de‐identified river drownings and 50 sea drownings were randomly selected from autopsy files at Forensic Science SA, Adelaide, Australia and assessed for decomposition. Results: The river cases consisted of 43 males and seven females (M:F 6:1); age range of 2–92 years (mean 46.8 years); decomposition—16/50 (32%); postmortem interval 1–14 days, average 3.8. The sea cases consisted of 36 males and 14 females (M:F 2.6:1); age range of 9–81 years (mean 50.7 years); decomposition—2/50 (4%) (p < 0.001%); postmortem interval 1–6 days, average 2.8. As decomposition reduces information obtained at autopsy, postmortem examination of deaths in rivers may be less informative than deaths that have occurred in the sea. The reasons for less decomposition in the sea include cooler temperatures, exposure to salt and shorter postmortem intervals.     

Glucose and lactate in vitreous humor compared with the determination of fructosamine for the postmortem diagnosis of diabetes mellitus

Diabetes mellitus is a chronic metabolic illness responsible for a great number of deaths. In postmortem diagnosis, because of the difficulty involved in interpreting blood glucose levels and relatively nonspecific pathologic features, biochemical markers in vitreous humor are useful. The aim of this study was to compare the results obtained for the combined determination of lactate and glucose with fructosamine levels recorded in the vitreous humor of two diagnostic groups (one diabetic and the other nondiabetic). The authors intended to ascertain the capacity of different markers measured in vitreous humor to diagnose diabetes mellitus. Fifty-one cadavers (mean age, 58.7 years; standard deviation, 17.09) were studied. The mean postmortem interval was 16.4 hours (standard deviation, 9.05). Cases were assigned to two diagnostic groups according to whether they were previously diagnosed as either diabetic or nondiabetic. Statistically significant differences for glucose, fructosamine, and the sum values of glucose and lactate were found between the two diagnostic groups. The highest levels were obtained in the group of cases with a previous diagnosis of diabetes mellitus. After the comparison of receiver operating characteristic curves, the sum values of glucose and lactate in vitreous humor is a better predictor of antemortem diabetes mellitus than the fructosamine.     

Melatonin concentrations in the sudden infant death syndrome

To examine a possible relationship between pineal function and the sudden infant death syndrome (SIDS), samples of whole blood, ventricular cerebrospinal fluid (CSF) and/or vitreous humor (VH) were obtained at autopsy from 68 infants (45 male, 23 female) whose deaths were attributed to either SIDS (n = 32, 0.5-5.0 months of age; mean +/- S.E.M., 2.6 +/- 0.2 months) or other causes (non-SIDS, n = 36, 0.3-8.0 months of age 4.3 +/- 0.3 months). The melatonin concentrations were measured by radioimmunoassay. A significant correlation was observed for melatonin levels in different body fluids from the same individual. After adjusting for age differences, CSF melatonin levels were significantly lower among the SIDS infants (91 +/- 29 pmol/l; n = 32) than among those dying of other causes (180 +/- 27; n = 35, P less than 0.05). A similar, but non-significant trend was also noted in blood (97 +/- 23, n = 30 vs. 144 +/- 22 pmol/l, n = 33) and vitreous humor (68 +/- 21, n = 10 vs. 81 +/- 17 pmol/l, n = 15). These differences do not appear to be explainable in terms of the interval between death and autopsy, gender, premortem infection or therapeutic measures instituted prior to death. Diminished melatonin production may be characteristic of SIDS and could represent an impairment in the maturation of physiologic circadian organization.     

Aminopeptidase and cathepsin A activity in vitreous humor in relation to causes of death

Brain autolysis happens rapidly, especially when environmental temperatures are high, and poses serious limitations for evaluating damage using morphologic methods. In the present study we have measured total proteins, cathepsin A and aminopeptidase activity in the vitreous humor in relation to cause of death and survival time. We have studied vitreous humor samples from 106 cadavers autopsied in the I.A.F. of Granada. The samples were classified according to causes of death as follows: myocardial infarction; hanging; other mechanical asphyxias; multiple trauma; craniocerebral trauma; other violent death; pulmonary functions; and other natural deaths. Total protein was measured by Lowry's method, and cathepsin A and aminopeptidase activity by the Bowen and Davison (1973) and Greenberg (1962) methods, respectively. Results are expressed in IU/l and in mIU/mg of protein. We found higher values of aminopeptidase and cathepsin A activity in groups with severe brain damage (craniocerebral trauma, multiple trauma, etc.) and lower values in groups of natural deaths. We believe, therefore, that aminopeptidase activity in vitreous humor may be a useful parameter for evaluating brain damage.     

Vitreous Humor: Biochemical Constituents in Estimation of Postmortem Interval*,†

Abstract: Analysis of biochemical constituents of the vitreous humor can be useful in determining the postmortem interval as there is proportionate postmortem rise of potassium and fall in sodium concentration. We studied 120 autopsy cases to determine the utility of potassium, sodium, calcium, and chloride levels, and sodium/potassium ratio in estimating the postmortem interval. There was a linear relationship between vitreous potassium concentration and postmortem interval, whereas an inverse relationship between vitreous sodium/potassium ratio and postmortem interval was noted. Other factors like age, sex, cause of death, season of death, and refrigeration of sample did not influence the vitreous humor potassium values. Using the statistical tools, a new formula was derived to determine the postmortem interval based on the potassium concentration and a review of previous literature is presented. Hence, the findings of this study supported a central role of vitreous humor biochemistry in many postmortem forensic and pathological evaluations.     

Vitreous humor carbohydrate-deficient transferrin concentrations in the postmortem diagnosis of alcoholism

Deaths from the effects of alcohol intoxication are encountered routinely in forensic practice. In an important number of cases difficulty may arise in interpreting the significance of results obtained in the autopsy. In clinical practice biochemical markers, particularly serum gamma-glutamyl-transpeptidase (GGT), alanine aminotransferase (ALT), aspartate transaminase (AST), carbohydrate-deficient transferrin (CDT), and erythrocyte mean corpuscular volume are used to diagnose heavy alcohol consumption. CDT is used as a reliable and specific marker. In postmortem diagnosis, because of the difficulty in interpreting blood alcohol levels and relatively non-specific pathological features, biochemical compounds have been studied for use as possible markers. The aim of this study was to evaluate the usefulness of the postmortem determination of CDT in vitreous humor as a confirmation of antemortem alcoholism. CDT levels were studied in 66 male cadavers with a mean age of 55.9 years (S.D. 17.0, range 22-87 years) with a mean postmortem interval of 17.9 h (S.D. 11.4, range 4-72 h). Cases were assigned to two diagnostic groups according to the antemortem diagnosis of alcoholism. Statistically significant differences were found for CDT and ALT concentrations between the two diagnostic groups. The highest vitreous humor levels of CDT and ALT were obtained in the group of cases with a previous diagnosis of alcoholism. Our results suggest that vitreous humor CDT levels are useful in cases where the postmortem diagnosis of alcoholism is hindered by the non-specificity of data.     

Basal renal tubular epithelial cell vacuolization and alcoholic ketoacidosis

Subnuclear renal tubular epithelial cell vacuolization is a marker for diabetic ketoacidosis. Whether it is because of hyperglycemia or of ketoacidosis is unclear. To examine the effect of ketoacidosis on renal cells in isolation, five cases of lethal alcoholic ketoacidosis without hyperglycemia were examined (vitreous humor β-hydroxybutyrate: 6.42-8.75 mM, mean 7.66 mM; and glucose: 0.1-4.2 mM, mean 1.46 mM). Microscopic examination of the kidneys revealed basal vacuoles in three cases (60%). Seven control cases with acute alcohol toxicity without ketoacidosis (blood alcohol: 0.18-0.43%, mean 0.31%; and β-hydroxybutyrate: 0.12-0.42 mM, mean 0.21 mM) did not have these changes. In this study, basal epithelial vacuolization was found only in cases with significant ketoacidosis. Although the numbers are small, the finding of basal renal tubular epithelial vacuolization in normoglycemic cases with elevated β-hydroxybutyrate levels provide further evidence that disordered lipid metabolism may be involved in the pathogenesis of this phenomenon.     

家兔急性胰腺炎死后不同时间眼玻璃体液淀粉酶变化

目的探讨急性胰腺炎死后不同时间眼玻璃体液淀粉酶变化。方法用1%牛胆酸钠诱导家兔急性胰腺炎动物模型建立,于死后不同时间和正常对照组,观察其淀粉酶变化。结果家兔急性胰腺炎死后72h眼玻璃体液淀粉酶含量(x)与死亡时间(y)存在相关关系,并导出其二项式回归方程为y=8.7420 0.7699x-0.0083x2(R2=92.62792,F=14.89734,P=0.001)。可作为推定早期损伤时间的参考指标。结论家兔急性胰腺炎死后不同时间眼玻璃体液淀粉酶改变为法医早期死亡时间推断提供了灵敏客观的实验依据。     

Diagnosis of sudden death in infants due to acute dehydration

When infants die suddenly of acute dehydration, clinical signs and autopsy findings may be equivocal or absent, and microbiologic cultures often are not helpful. Vitreous humor electrolyte and urea nitrogen concentrations were measured in 53 infants dying of gastrointestinal infections, sudden infant death syndrome (SIDS), other infectious diseases, and miscellaneous causes to determine whether these parameters would assist in the recognition and confirmation of deaths resulting from dehydration. Significant differences were found when comparing the mean sodium and urea nitrogen levels of infants dying of gastrointestinal infections with those succumbing to SIDS or other causes. We recommend that these determinations be routinely performed whenever the gross autopsy findings are insufficient to explain the death.     

Postmortem vitreous humor beta-hydroxybutyrate: its utility for the postmortem interpretation of diabetes mellitus

Ketoacidotic coma is one of the most serious complications arising from diabetes mellitus, especially type I, and may be the cause of sudden death especially in diabetes type I. Since beta-hydroxybutyrate (beta-OHB) serum concentrations might provide more information on the severity of ketoacidosis, the aim of this study was to evaluate the concentrations of beta-OHB in vitreous humor and its correlation with other biochemical parameters during postmortem examination. We intended to ascertain the sensitivity and the specificity of these markers for diagnosing diabetes mellitus and the presence of ketoacidosis. This study involved 453 cadavers with a mean age of 57.6 years (S.D. 20.7) and a mean postmortem interval of 17.8 h (S.D. 9.6, range 2-61 h). Cases were assigned to two diagnostic groups according to the antemortem diagnosis of diabetes mellitus, based on the patients' medical records. In vitreous humor statistically significant differences were found in biochemical marker concentrations between the two diagnostic groups, the highest values being obtained in the group of subjects with a previous diagnosis of diabetes mellitus. The measurement of beta-OHB in vitreous humor may be a useful alternative to using blood during postmortem analysis. The presence of high levels of beta-OHB may help interpret the cause of death in diabetics when the autopsy result is negative.     

Obesity,Body Mass Index,and Homicide

The body mass indexes (BMIs) of 100 randomly selected homicide cases from the files of Forensic Science SA were compared to the Australian and South Australian populations. There were 70 males and 30 females (M:F = 2.3:1; age range 18–84 years; mean 42.3 years). There was a substantially lower proportion of obese individuals in the homicide population compared to the general Australian and South Australian populations (19% [vs.] 27.9% and 30%, respectively). A second group of 144 randomly selected autopsy cases where the BMI was ≥40 kg/m² was analyzed. There were 77 males and 67 females (M:F = 1.2:1; age range 23–78 years; mean 46.7 years). The majority of deaths were natural (N = 108), with no homicides. A negative association between obesity and homicide has, therefore, been demonstrated. Reasons for the lower numbers of obese/morbidly obese individuals among homicide victims are unclear, but may include physical protection afforded by fat padding from sharp force injuries, and relative sociodemographic isolation.     

家兔急性有机磷中毒死后玻璃体液化学成分的变化趋势

目的 探讨家兔急性有机磷中毒死后不同时间眼玻璃体液化学成分的变化。方法用5％敌百虫制作家兔急性有机磷中毒动物模型,在死后0～72h内不同时间点,采用留置微量重复取样法吸取眼玻璃体液,以全自动生化分析仪测量眼玻璃体液中钾、镁、磷、钙、钠、氯浓度的变化,统计数据进行二项式回归方程拟合分析。结果家兔急性有机磷中毒死后72h眼玻璃体液中钾、镁、磷随PMI延长而逐渐升高,舻分别为0．9818、0．9700、0．9458,与PMI相关性好,其中30h内镁的相关性更好;钙、钠随PMI延长呈降低趋势,砰分别为0．7124、0．7355,相关性较差;氯表现不稳定掣仅为0．0869。结论家兔急性有机磷中毒死后玻璃体液中钾、镁、磷、钙、钠等离子浓度随PMI延长而改变,72h内总体衡量,钾与PMI最具有相关关系。     

References for determining the time of death by potassium in vitreous humor

The different statements concerning the slope and intercept of the regression line and the 95% limits of confidence are the reason that potassium in vitreous humor is not used (at least in Germany) as an aid in estimating the time of death. The relationship between the concentration of potassium and the time of death is mainly influenced by antemortem electrolyte imbalances caused by disease and/or duration of terminal episode. The influence of terminal episode is best identified by its duration (Adelson et al., J. Forensic Sci., 8 (1963) 503–514). In order to have a method suitable for every case and to be as precise as possible we looked therefore for parameters in vitreous humor which were stable postmortem and indicating antemortem electrolyte imbalance. Urea is such a parameter, being stable postmortem (Coe, Am. J. Clin. Pathol., 51 (1969) 741–750) and useful as a marker of antemortem electrolyte imbalance. Our investigations on potassium in vitreous humor, including sudden and hospital deaths after chronic lingering disease, revealed 95% limits of confidence of ±34 h up to 120 h postmortem. Reviewing only cases with urea less than 100 mg/dl the 95% limits of confidence could be reduced to ±22 h. Considering the duration of terminal episode (<6 h) the precision was ±20 h. In this way our modified procedure is suitable for every case with the resulting precision of estimation being determined only by the duration of the terminal episode and urea concentration.     

Comparison of chemical methods for determining postmortem interval

Accurate determination of postmortem interval (PMI) is a problem for the forensic thanatologist, especially in unwitnessed deaths. A number of objective chemical methods for determining PMI have been developed, the most widely used being accumulation of potassium in the vitreous humor. The authors previously have reported a chemical method for determining PMI from the predictable accumulation or clearance of the dopaminergic metabolite 3-methoxytyramine (3-MT) in the putamen of the brain. They have extended their previous study to compare directly the accuracy of determining PMI from the level of 3-MT in putamen with the level of potassium in vitreous humor. The data indicate that 3-MT is at least as accurate as, if not more accurate than, potassium accumulation in vitreous humor, although 3-MT levels can be affected by the cause of death and drugs present at the time of death. Nevertheless, determination of both the 3-MT and potassium levels can afford the most accurate method of determining PMI; preliminary nomograms for determining PMI from both variables are presented.     

References for determining the time of death by potassium in vitreous humor

The different statements concerning the slope and intercept of the regression line and the 95% limits of confidence are the reason that potassium in vitreous humor is not used (at least in Germany) as an aid in estimating the time of death. The relationship between the concentration of potassium and the time of death is mainly influenced by antemortem electrolyte imbalances caused by disease and/or duration of terminal episode. The influence of terminal episode is best identified by its duration (Adelson et al., J. Forensic Sci., 8 (1963) 503-514). In order to have a method suitable for every case and to be as precise as possible we looked therefore for parameters in vitreous humor which were stable postmortem and indicating antemortem electrolyte imbalance. Urea is such a parameter, being stable postmortem (Coe, Am. J. Clin. Pathol, 51 (1969) 741-750) and useful as a marker of antemortem electrolyte imbalance. Our investigations on potassium in vitreous humor, including sudden and hospital deaths after chronic lingering disease, revealed 95% limits of confidence of +/- 34 h up to 120 h postmortem. Reviewing only cases with urea less than 100 mg/dl the 95% limits of confidence could be reduced to +/- 22 h. Considering the duration of terminal episode (less than 6 h) the precision was +/- 20 h. In this way our modified procedure is suitable for every case with the resulting precision of estimation being determined only by the duration of the terminal episode and urea concentration.     

Ethyl sulphate and ethyl glucuronide in vitreous humor as postmortem evidence marker for ethanol consumption prior to death

To clarify the circumstances of death, the degree of inebriation is of importance in many cases, but for several reasons the determination of the ethanol concentration in post-mortem samples can be challenging and the synopsis of ethanol and the direct consumption markers ethyl glucuronide (EtG) and ethyl sulphate (EtS) has proved to be useful. The use of a rather stable matrix like vitreous humor offers further advantages. The aim of this study was to determine the concentrations of ethanol and the biomarkers in the robust matrix of vitreous humor and to compare them with the respective levels in peripheral venous blood and urine. Samples of urine, blood from the femoral vein and vitreous humor were taken from 26 deceased with suspected ethanol consumption prior to death and analyzed for ethanol, EtS and EtG. In the urine samples creatinine was also determined. The personal data, the circumstances of death, the post-mortem interval and the information about ethanol consumption prior to death were recorded. EtG and EtS analysis in urine was performed by LC-ESI-MS/MS, creatinine concentration was determined using the Jaffé reaction and ethanol was detected by HS-GC-FID and by an ADH-based method. In general, the highest concentrations of the analytes were found in urine and showed statistical significance. The mean concentrations of EtG were 62.8mg/L (EtG100 206.5mg/L) in urine, 4.3mg/L in blood and 2.1mg/L in vitreous humor. EtS was found in the following mean concentrations: 54.6mg/L in urine (EtS100 123.1mg/L), 1.8mg/L in blood and 0.9mg/L in vitreous humor. Ethanol was detected in more vitreous humor samples (mean concentration 2.0g/kg) than in blood and urine (mean concentration 1.6g/kg and 2.1g/kg respectively). There was no correlation between the ethanol and the marker concentrations and no statistical conclusions could be drawn between the markers and matrices.     

Hemolytic staining of the intima of the aortic root in freshwater drowning: a retrospective study

A retrospective study was conducted of files at the Institute of Legal Medicine in Hamburg, Germany from 1998 to 2003 of all cases of drowning where there was no putrefaction, to document the rate of recording of hemolytic staining of the intima of the great vessels. All cases had full police investigations with reviews of previous histories, circumstantial evidence, and autopsy findings. A series of control subjects who had died of various natural and nonnatural, nondrowning deaths, matched for age, sex, and postmortem interval, was also reviewed. One hundred twenty cases of freshwater drowning were identified. Drowning occurred in a garden pool in 1 case, in a bath in 2, and in the river Elbe in 117. The age range was from 2 to 91 years (mean = 55 years; M:F = 1:1.8). Hemolytic intimal staining of the aortic root was documented in 6 cases (6 of 120; 5%). This consisted of reddened discoloration of the proximal portion of the aortic root, without any significant staining of the proximal pulmonary artery. No significant hemolytic staining of the intima of the great vessels was recorded in any of the 120 control cases. Although under-reporting of findings may occur in retrospective analyses, this study has shown that at least 5% of freshwater drowning cases showed differential staining of the pulmonary trunk and aorta, with hemolytic discoloration of the aortic intima. When present, hemolytic staining of the aortic root intima may be a useful and possibly under-recognized corroborative sign of freshwater drowning.     

Vitreous humor magnesium in alcoholics

Postmortem vitreous humor magnesium concentrations were determined in 27 alcoholics and in 27 nonalcoholics who died of various causes. No significant differences in magnesium values were present between these two groups. The concentrations in those dying with alcohol withdrawal syndrome or in those with postmortem findings limited to hepatic fatty change did not differ significantly from those in the nonalcoholic group or from those of the alcoholics dying of other causes. There was no correlation between vitreous humor magnesium concentrations and postmortem interval in any group. We conclude that vitreous humor magnesium determination is not helpful in the postmortem evaluation of suspected alcohol-related deaths or in determining postmortem interval.     

Subnuclear renal tubular vacuoles in alcohol use disorder

Subnuclear vacuoles in the proximal renal tubules have been reported as a histologic sign of ketoacidosis. Originally described in diabetic ketoacidosis, renal vacuoles can be found in other ketogenic states such as alcoholic ketoacidosis (AKA), starvation, and hypothermia, underpinned by deranged fatty acid metabolism. A retrospective analysis of 133 deaths associated with alcohol use disorder (AUD) examined at autopsy between 2017 and 2020 was undertaken. This study aimed to determine the prevalence of subnuclear vacuoles in deaths of those with AUD and their specificity for deaths from AKA, and to elucidate what demographic, biochemical, and pathologic findings are associated with subnuclear vacuoles. In each case, vitreous humor biochemistry including electrolytes, glucose, and beta-hydroxybutyrate (BHB) was analyzed alongside postmortem hemoglobin A1c and renal and liver histology. Renal histology was graded for the presence of vacuoles as absent (0), scanty (1), or easily identifiable (2). Liver histology was graded for steatosis and for fibrosis if Masson trichrome staining was available. Vacuoles were commonly seen in the deaths of those with AUD. They were seen in deaths due to AKA but were not specific to that cause of death. With vacuoles present, lower vitreous sodium (139 vs. 142 mmol/L; p = 0.005), higher vitreous BHB (1.50 vs. 1.39 mmol/L; p = 0.04), severe hepatic steatosis, and severe hepatic fibrosis were seen, compared with those without renal vacuoles.     

Vitreous humor biochemical constituents: evaluation of between-eye differences

The present study was conducted to investigate the differences in the vitreous humor biochemical concentrations for vitreous electrolytes and calcium in the same pair of eyes at identical postmortem interval (PMI). The vitreous humor samples were collected independently in both eyes from 48 autopsies (PMI range, 4.5-84.3 hours) with documented time of death. The samples were analyzed for potassium, sodium, chloride, and calcium using a Beckman Coulter LX20 Automated Analyzer based on ion-selective electrode methodology. There were no statistically significant between-eye differences at identical postmortem interval. A significantly high correlation was observed between paired potassium concentrations of both the eyes. A highly significant linear correlation was observed between the individual eye and mean potassium concentrations of both the eyes with postmortem interval. The observed differences were not significantly correlated with postmortem interval. The results demonstrated that the between-eye differences for vitreous electrolytes and calcium are insignificant. Therefore, the utility of vitreous biochemistry, particularly potassium in postmortem interval estimation and other forensic applications, cannot be questioned solely on the basis of these differences.