溺死诊断中的浮游生物检测法

溺水是常见的死亡原因之一,浮游生物检验是法医学鉴定溺死的重要依据。浮游生物检验方法可分为破机法、非破机法和分子生物学方法三大类。前两者主要以形态标记为检验对象,依赖于浮游生物的理化特性,有很大局限性。分子生物学方法以分子标记为检验对象,适用范围广,信息丰富,是值得探索的新方法。本文回顾溺死鉴定中的各种浮游生物检测法,供法医工作者在实践和研究中进行参考。     

浮游生物PCR检测及其在溺死鉴定中的研究进展

溺死鉴定是法医学中的重点和难点之一,目前在相关法医学案件中,主要采用法医病理学尸检与传统硅藻检验等方法综合分析进行鉴定。传统硅藻检验存在灵敏性较低,易于污染等不足。采用PCR法检测尸体不同脏器组织中是否存在水中浮游生物的DNA标记,适用范围广,信息量丰富,灵敏性和特异性较高,有较好的法医学应用前景,有望成为鉴定溺死的新方法。本文对相关研究进展进行综述,为相关研究和实践参考。     

鉴定溺死的实验研究——荧光分光光度计检测肺组织浮游生物叶绿素(a)

本实验应用荧光分光光度计检测肺组织浮游生物叶绿素(a),以期通过准确的定性、定量研究来诊断溺死。实验性研究表明本法准确可靠,选择性强、灵敏度高,不受腐败及污染等因素影响,亦不存在含叶绿素(a)浮游生物生前进入并存在于人体器官的问题。同法对2例水中尸体进行了检测,结果与实验性研究相符、提示本法可应用于实际检案。     

Postmortem chest computed tomography for the diagnosis of drowning:a feasibility study

It may be difficult to distinguish the cause of death in drowning cases without specific findings. The aim of this study was to explore the forensic value of thoracic postmortem computed tomography (PMCT) using routine images and three-dimensional (3D) image reconstructions. The imaging data of PMCT examinations of six drowning cadavers, aged 21–54 years, were analyzed. Twelve victims of sudden death from coronary artery disease (CAD) were chosen as a control group. After 3D bilateral lung images were reconstructed using image processing software, an interactive medical image control system was used to measure and analyze parameters including lung volume, lung volume ratio, mean CT value of the whole lung, and lung CT value distribution curves. Lung volume and lung volume ratio were used to assess the shape changes of the lung. Lung CT value distribution curves showed the corresponding number of pixels of the different CT values in the lung image. Lung volume was not significantly larger in drowning cases (mean 2 958 cm³) than in controls (mean 2 342 cm³). Lung volume ratio values in the drowning group (mean 0.3156) were greater than those in the control group (mean 0.2763); (P = 0.02). There was no significant difference between the drowning and control group in the mean CT value of the whole lung. There were differences between lung CT value distribution curves in drowning victims and controls, with drowning victims showing a single peak and CAD cases showing a bimodal distribution. Thoracic PMCT is helpful for the forensic medical diagnosis of drowning. Lung volume ratio and lung CT value distribution are potential indicators to distinguish between drowning and CAD.     

浮游生物16S rDNA检测在大鼠溺死鉴定中的价值

目的建立浮游生物16S rDNA PCR检脸法应用于溺死的鉴定。方法将15只SD大鼠随机分为溺死组、死后抛尸组和正常对照组,死后分别取脑、肝、肾和肺等脏器,提取DNA.用特异性引物扩增浮游生物16S rDNA第三和第四可变区特异片段。结果溺死组5例,肺全部检出(100%)浮游生物16S rDNA;肝、肾分别检出4例(80%);脑检出3例(75%)。死后抛尸组仅1例在肺中检出浮游生物(20%)。正常对照组均未检出。苗论浮游生物16S rDNA PCR检验法可用于溺死的鉴定。     

The usefulness of lung surfactant phospholipids (LSPs) in the diagnosis of drowning

The authors have studied the usefulness of some lung surfactant phospholipids (LSPs) isolated from lung tissues as markers of drowning. Two different groups of rabbits were sacrificed by drowning in fresh and salt water, and their phospholipid compositions were compared with those of a non-drowned control series. For the phospholipids studied in lung lavages (phosphatidyl choline, phosphatidyl ethanolamine, and phosphatidyl glycerol) the proportions differed between the control group and the drowned group, and between the fresh-water and salt-water drowned animals. According to these results, the lipids we have analyzed can be employed as markers in forensic autopsies, where it is necessary to differentiate between death by drowning and postmortem immersion and between fresh-water and salt-water drowning. In lung tissue, only phosphatidyl choline and phosphatidyl inositol showed significative differences. These results also confirm that LSPs are strongly affected in drowning.     

大鼠溺死后肺组织与血清IL-1β、IL-13mRNA表达变化

目的检测大鼠肺组织和血清中IL-1β、1L-13mRNA表达变化,探讨IL-1β、IL-13mRNA的变化情况与溺死的关系。方法 12只SD大鼠随机分为对照组、溺死组(实验组)。分别提取各组大鼠右侧肺下叶及右心室血清,应用TaqMan探针法检测肺组织及右心室血清IL-1β、IL-13mRNA的表达情况。结果 (1)溺死组大鼠肉眼观察:体表征象及解剖所见符合生前溺死特征。(2)肺组织中IL-1β、IL-13mRNA表达检测:与对照组比较,实验组IL-1β、IL-13表达呈下降趋势,差异无统计学意义(p0.05)。(3)血清中IL-1β、IL-13mRNA表达检测:与对照组比较,实验组IL-1β表达显著升高(p0.01)、IL-13表达显著升高(p0.01)。结论 (1)IL-1β、IL-13 mRNA在溺死组肺组织中表达呈下降趋势,可能是大鼠溺死出现代偿性抗炎反应综合症导致免疫无能的表现。(2)IL-1β、IL-13 mRNA在血清中表达明显升高,可能与溺水应激及溺死后急性肺损伤创伤应激有关。     

PCR-DHPLC法检测硅藻SSU基因在溺死鉴定中的应用

目的采用PCR-DHPLC法检测硅藻SSU基因,评估其在溺死鉴定中的应用价值。方法 60只实验兔随机分为生前溺死（水中溺毙）、死后入水（空气栓塞致死后入水）、对照组（空气栓塞致死后不做处理）;溺死人体脏器组织;取各组织检材提取硅藻DNA,PCR扩增硅藻特异的核糖体小亚基（SSU）片段,用琼脂糖凝胶电泳检测、DHPLC检测分析。结果 6份硝酸消化法检测阴性的溺死人体器官组织检材经PCR及琼脂糖凝胶电泳检出5例阳性。生前溺死组肺、肝、肾硅藻检出率分别为100%、90%、85%,死后入水组仅肺检出硅藻（15%）,对照组各组织均为阴性;生前溺死组检出率明显高于死后入水组（P〈0.05）。10份溺死人体器官组织检材采用DHPLC法检出硅藻种类明显多于微波消解-扫描电镜法（P〈0.05）。脏器检出硅藻种类与溺死点水样一致。结论采用PCR-DHPLC法检测硅藻SSU基因,有助于溺死鉴定和溺死地点的推断,具有法医学应用价值。     

Complex Suicide by Self‐stabbing and Drowning: A Case Report and a Review of Literature

Sharp force injuries and drowning are methods exceptionally combined in complex suicides. We report a challenging case of complex suicide by self‐stabbing and drowning that illustrates the difficulty in discriminating between homicide and suicide in such circumstances of death. The corpse of a young man was found submerged in a river, stabbed nine times with two wounds that had penetrated the thorax and had caused lung injuries and a hemopneumothorax. The postmortem and histological examinations were consistent with a death caused by drowning, but the manner of death still remained undetermined. Police investigation finally concluded to a suicide, although no suicide note had been left and the victim had no underlying diagnosed mental disorder. The parameters that may help distinguish suicide from homicide at the autopsy should be interpreted in light of a thorough forensic investigation to determine the exact manner of death in such a case.     

Comparison between lung weight and blood strontium in bodies found in seawater

This paper examines the use of lung weight increase as an indicator of seawater drowning compared to the amount of Sr absorbed by the blood. The study population was limited to male victims older than 20 years. Significant differences between cases of drowning and "non-drowning" were detected in terms of the lung-heart weight ratio (L/H) (p<0.001) or lung-body weight ratio (L/B) (p=0.005). However, using lung weight (L), L/H or L/B to distinguish between seawater drownings and saltwater non-drownings some overlap was produced. The factor rendering least overlap was L/B, which also appeared to be non-dependent on the victim's age. Our findings suggest that a value of L/B higher than 19.5 g/kg could be a useful indicator of death by drowning, but that when a lower value is found, additional drowning diagnoses would be needed to establish the manner of death.     

硅藻定量检验和种属鉴定的实验研究

本实验用家兔18只,分为溺死组、死后(抛尸)入水组及对照组,每组各6只。采用消化—光镜法作肺、肝、肾硅藻计数(定量)及种属鉴定检验,同时作水样检验及空白对照。结果:(一)进一步证实生前硅藻可经空气源性进入肺脏并存积于大循环内脏(肝、肾)的概念;(二)提示水中尸体肺脏如有大量硅藻检出,仅此即可作为诊断溺死的可靠依据;空气源性吸入及检验过程中的污染并不影响该检验结果对鉴定溺死的判断。种属分析溺死组肺与水样的主要硅藻种属相符,而肝、肾中硅藻计数及种属分析均无诊断意义。     

非溺死尸体肺脏硅藻最大值在溺死鉴定中的应用初探

探讨非溺死尸体肺脏硅藻最大值（即空气吸入与检验污染之和的最大值）在溺死鉴定中的应用价值。用硝化-光镜法检测贵阳地区40例非溺死尸体肺脏硅藻含量的最大值，同时作了8例溺死尸体肺脏硅藻检验。非溺死组肺脏硅藻最大值2个／5g；溺死组有7例为56～463个／5g，1例是2个／5g。表明（1）水中尸体肺脏硅藻检验值大于最大值（2个／5g），可考虑为溺死；显著大于最大值（本文溺死组7例56～463个／5g）则可鉴定为溺死；（2）等于或小于最大值，提示干性溺死或抛尸入水，可结合尸检发现及案情鉴别。     

Selective culturing and genus-specific PCR detection for identification of Aeromonas in tissue samples to assist the medico-legal diagnosis of death by drowning

The detection of autochthonous aquatic bacteria in tissue samples from drowning cases is increasingly considered as an alternative approach to assist the medico-legal diagnosis of death by drowning. Bacteria belonging to the genus Aeromonas may be suitable candidates for this application as they are ubiquitous in natural aquatic environments but are generally not part of the human microbiota. The research aims of this study were (i) to develop a sensitive, specific and rapid screening and confirmation method for Aeromonas species in tissue samples and (ii) to evaluate aseptic sternal puncture as a post-mortem sample technique and bone marrow as an alternative matrix to provide evidence of death by drowning. The presence of Aeromonas in tissue samples was verified by cultivation using the selective media Ampicillin Dextrin Agar (ADA) and Ryan's Aeromonas Medium. The use of ADA medium was found most optimal for the sensitive, inexpensive and quick detection of aeromonads in human tissue samples. Positive culture plates were confirmed by harvesting all colonies for DNA extraction and subsequent PCR amplification using Aeromonas genus-specific primers. Aeromonads were detected in lung swab, blood and bone marrow of drowned bodies (n=3), but were negative in these three matrices for all negative controls (n=90) tested. Bone marrow proved to be a suitable alternative matrix and can be sampled post-mortem by an aseptic sternal puncture. In conclusion, this study confirms previous indications that aeromonads in cultures from blood of water bodies can be considered a potential marker for drowning. Given the fact that the number of immersed bodies (drowned and non-drowned) included in this study is statistically not significant, however, more tissue samples need to be investigated to confirm the validity of these methods to aid the diagnosis of death by wet drowning.     

Organ weight effects of drowning and asphyxiation on the lungs, liver, brain, heart, kidneys, and spleen

An examination of the organ weights associated with victims of drowning, asphyxiation and trauma was undertaken to determine (a) the effects of asphyxiation compared to a trauma group, and in turn, (b) the effects of drowning compared to an asphyxiation group. Included in the study were 217 drowning deaths, 166 pure asphyxiation deaths and 381 trauma deaths. The effects of asphyxiation (compared to trauma) resulted in elevated mean organ weights for the lungs, liver, kidneys and spleen (with mean increases of 17.8, 10.5, 10.3 and 23.4%, respectively). Effects of drowning (compared to asphyxiation) resulted in elevated mean organ weights only with the lungs and kidneys (with mean increases of 30.0 and 4.4%, respectively). Only the mean heart and brain weight remained constant across all experimental groups. A picture of drowning is suggested in which elevated lung and kidney weights are the result of both asphyxiation and the aspiration of water that occurs with drowning, whereas elevated spleen and liver weights in drowning victims are associated with only the effects of asphyxiation. In addition, the common autopsy finding of a small, anemic spleen in drowning, rather than caused by some pathophysiological mechanism of death, is hypothesized to be a postmortem phenomenon.     

Diagnostic significance of myofibrillar degeneration of cardiocytes in forensic pathology

The incidence of myofibrillar degeneration (MFD) was studied in the following different forensic-pathological diagnostic groups of 25 cases each: acute morphine intoxication, acute carbon monoxide intoxication, hanging, strangulation by hand/ligature, drowning, acute hemorrhagic shock, lethal acute brain injury, explainable death of babies or infants and sudden infant death syndrome, together with 18 cases of intoxication with various drugs. The MFD was demonstrated by the Luxol-fast-blue reaction, with two types of phenomena being differentiated, namely cross-band lesions and diffuse staining. All diagnostic groups included cases of MFD of differing degrees. Cross-band lesions were observed in practically all cases of hanging, strangulation and acute hemorrhagic shock. Diffuse stain was noted particularly in cases of drowning and acute brain injury. The diagnostic significance is discussed.     

Organ weight effects of drowning and asphyxiation on the lungs,liver, brain,heart, kidneys,and spleen

An examination of the organ weights associated with victims of drowning, asphyxiation and trauma was undertaken to determine (a) the effects of asphyxiation compared to a trauma group, and in turn, (b) the effects of drowning compared to an asphyxiation group. Included in the study were 217 drowning deaths, 166 pure asphyxiation deaths and 381 trauma deaths. The effects of asphyxiation (compared to trauma) resulted in elevated mean organ weights for the lungs, liver, kidneys and spleen (with mean increases of 17.8, 10.5, 10.3 and 23.4%, respectively). Effects of drowning (compared to asphyxiation) resulted in elevated mean organ weights only with the lungs and kidneys (with mean increases of 30.0 and 4.4%, respectively). Only the mean heart and brain weight remained constant across all experimental groups. A picture of drowning is suggested in which elevated lung and kidney weights are the result of both asphyxiation and the aspiration of water that occurs with drowning, whereas elevated spleen and liver weights in drowning victims are associated with only the effects of asphyxiation. In addition, the common autopsy finding of a small, anemic spleen in drowning, rather than caused by some pathophysiological mechanism of death, is hypothesized to be a postmortem phenomenon.     

Morphologic determinants of asphyxia in lungs: a semiquantitative study in forensic autopsies

Asphyxia is a name given to different kinds of lesions that can produce similar histologic findings. Thus, because of the varied nature of the different kinds of lesions, as well as the incidence of similar qualitative histologic findings with different causes, the aim of this work was to study special kinds of injuries with particular subsequent impairment. These include some diagnostic problems of sudden death of natural causes, including aspiration, suffocation, drowning, and strangulation. Ranking was made of 167 victims based on the diagnosis as having: aspiration (n = 35), suffocation (n = 88), drowning (n = 27), and strangulation (n = 17). Stepwise discriminant analysis of the resulting data showed that lung necropsies from victims of these four events could be distinguished from one another. Statistical differences among the four groups were observed for eight morphologic parameters. A robust discriminant function permitted an adequate classification of the four groups of disease in 85.03% of the cases. Lung autopsies with congestion, septal hemorrhage, and foreign body showed a specificity of 100% for victims of aspiration, whereas ductal overinsufflation, interstitial edema, and bronchiolar constriction showed a specificity of 81.8% in victims of suffocation. Intraalveolar edema and dilatation of the alveolar spaces with secondary compression of the septal capillaries characterized drowning. Victims of strangulation showed a strong alveolar hemorrhage, with alveolar collapse and overinsufflation, associated with bronchiolar dilatation. It is concluded that semiquantitative analysis of lung autopsies might be a useful supplementary histologic criterion to support the diagnosis of asphyxia.     

检测浮游生物叶绿素相关基因诊断溺死的实验研究

目的评估浮游生物叶绿素相关基因检测用于溺死诊断的价值。方法将18只大白兔随机分为溺死组（n=10）、死后抛尸组（n=6）和对照组（n=2）,各组分别取心血及肺、肝、肾、脑组织,分离浮游生物并提取其DNA,用PCR技术检测叶绿素相关基因EG（EG1和EG2）及SK（SK1和SK2）。同时用硝酸消化法检验肺和肝组织中的硅藻。结果溺死组心血及肺、肝、肾、脑组织中EG1分别检出9、10、9、7和8例阳性,EG2分别检出8、10、7、5和7例阳性;死后抛尸组仅在心血和肺组织中各检出1例EG1阳性;对照组各组织均未检出EG1和EG2。SK1、SK2除在溺死组心血、肺和肾有少数检出外（≤2例）,在其他组未检出扩增产物。硝酸消化法从溺死组肺、肝组织中分别有9例及3例检出硅藻,死后抛尸组仅在1例肺组织中检出。结论浮游生物叶绿素相关基因EG用于溺死诊断的阳性检出率要高于硝酸消化法,在溺死诊断中具有较高的应用价值。     

Pediatric drowning: a 20-year review of autopsied cases: II. Pathologic features

The pathologic findings in autopsies of drowning victims are nonspecific and vary from case to case. However, most reported pathologic series of drowning cases exclude children and do not take into consideration the unique circumstances surrounding bathtub drownings. In addition, the effect of resuscitation on the autopsy findings has not been studied in children. A retrospective review of autopsy records of non-bathtub drownings from a 20-year period (1984-2003) was performed and 63 cases were identified in 45 males and 18 females (age range 9 months to 17 years). The incidence of frothy exudate, pleural effusion, and increased lung weight was 43%, 36%, and 80%, respectively. The incidence of frothy exudate and the combination of all 3 factors was significantly higher in cases with no resuscitation compared with those cases with attempted resuscitation with or without delayed death. As the interval between the drowning episode and autopsy increased, the incidence of frothy exudate decreased significantly. There was no relationship between these findings and the age and sex of the decedent. Other clinical conditions or occult pathologic findings that may have contributed to death were found in 8 cases (13%). The findings highlight the need for thorough clinicopathologic correlation in cases of drowning to accurately interpret the pathologic findings.     

The development and application of a diatom-based quantitative reconstruction technique in forensic science

Diatoms are a group of unicellular algae that have been recorded and classified for over 200 years and have been used in a range of applications in forensic science. We have developed a quantitative diatom-based reconstruction technique to confirm drowning as a cause of death and localize the site of drowning in two recent, high-profile, case studies. In both case studies we collected diatom samples from the local and/or regional area to act as a control in the examination of diatom assemblages associated with lungs and clothing. In Case Study 1 the modern analog technique suggested that all lung and clothing samples have statistically significant similarities to control samples from shallow water habitats. In Case Study 2, the analog matching suggested that the majority of lung samples show a statistically significant relationship to samples from a pond, indicating that this was the drowning medium.