| Object:Drowning is the liquid mechanically blocked airway and prevented gas exchange, so that the body lack of oxygen, and the carbon dioxide retention, leading to death. The coastal areas have abundant cases of drowning, so that the forensic autopsy of bodies in water is an important part of works for forensic scientists. Drowning is the most cause of death in routine cases in water bodies, however, but there are some situations of possible things that bodies were died before been thrown into the water. Consequently, the bodies in the water may be due to criminal conducts, and must be conducted in forensic autopsy and related test. Drowned corpses may emerge some special pathological signs, such as the mushroomed foam in mouth and nose, aqueous emphysema, and exogenous objects in the bronchial airway. Some drowned bodies and deep corruption corpses are without those special pathological signs, however, should employ some professional measures to assist diagnosis of drowning. There are a lot of methods to diagnose drowning, but all of these methods have some defects that lack of specificity and affected by bodies’ corruption and restriction of technology, so that these methods are difficult to apply to forensic drowning diagnosis. Diatoms in drowning medium are inhaled with severe breathing and get through alveolar capillary barrier into the blood vessels, and arriving organs in the bigger circulation finally, in the procession of drowning. The procession combines with the characteristic of diatoms’ wide range in the water make diatom test can assist diagnosis of drowning.At present, the diatom test is considered to be the effective measure in diagnosis of drowning of the bodies in water, especially in bodies of deep corruption. Diatom test practice mainly consists of three processions:digestion, enrichment and detection. Many methods were used that were few advantages, leading to exist some problems. Such as traditional acid digestion method characters simple procedure, low cost, but lower enrichment efficiency and sensitivity, and scanning electron microscopy(SEM) detected diatoms characters easy observation and been taxon, but its high cost limited its applications. This paper aims to establish a new methods characters effective rate of detection, simple, safe, easy equipments and low cost in diatom test. It can get great effects in the course of diatom test, and provide a suitable measure for forensic science laboratories.Methods:The development of method:alcohols, aldehydes, acids, benzene, resin and other reagents and membrane of the polyether sulfone (PES) material, membrane of the nylon material and the nuclear pore membrane (organic material) were tested to explore and establish a new method of diatom test based on membrane enrichment and optical microscopy from organs and water. And its’features was discussed too.Animal experiments:10 New Zealand white rabbits were killed ethically, and kidneys were extracted.500ml water samples in the Pearl River of the Guangzhou were sampled too. Samples were randomly divided into strong acid digestion-centrifugal enrichment-light microscopy group (group A), microwave digestion-membrane enrichment-scan electric microscopy group (group B), and microwave digestion-membrane enrichment-light microscopy group (group C). Each group has 20 samples, and were used strong acid digestion-centrifuge enrichment-light microscopy method (method A), microwave digestion-membrane enrichment-SEM method (method B) and method established by this study that microwave digestion-membrane enrichment-light microscopy (method C), respectively. Test indexes were sampling time, the extent of digestion, the total recovery rate of diatoms, each diatom species recovery rate, and application.Cases test:56 Bodies in the water were collected from February 2013 to October 2013, Guangzhou City. It were collected that 30 bodies dead of drowning, and 26 bodies dead of murder and throw into water, and 30 bodies dead of diseases other than drowning, Guangzhou urban area.lungs, livers, kidneys and control water samples were taken and were conducted by strong acid digestion-centrifuge enrichment-light microscopy method (method A), microwave digestion-membrane enrichment-SEM method (method B) and microwave digestion-membrane enrichment-light microscopy method (method C). Test indexes were sampling time, observation effect and application.Case application:Six cases were used microwave digestion-membrane enrichment-light microscopy, which collected from December 2013 to April 2014. Cases’ lungs, livers, kidneys tissue and the control water were imposed by diatom test. Test indexes were diatom numbers and species detected in samples and the results of these cases were tracked, too.Results:Development of the method:eugenol- acetic acid mixed reagent can make membrane transparent immediately. Forensic scientists can observe the sample directly under an optical microscope, and the diatoms can be detected clearly and easily. PES membranes and eugenol- acetic acid mixed reagent are suitable for this study, furthermore, a new method of diatom test was established:microwave digestion-membrane enrichment-light microscopy.Animal experiments:The time of sampling in three groups were shown that the method A was the longest, and the time of method B was longer than Method C, which was the shortest one (P<0.05). The digestion ability of organs in Method A was incomplete, which there were lots of massive tissue surrounding diatoms under the light microscope, even some diatoms were covered by parts of incomplete tissue, leading to difficult observed. The tissues were digested completely under Method B. There were no residues under the light microscope, and the textures of diatoms were observed clearly, leading to be counted and identified species easily. The tissues were digested completely under method C, too. Thus simples were good transparency, and diatoms were easy to be counted and identified under the light microscope. The diatoms total recoveries of the three methods were 2.6±1.7%,76.9 ±20.2% and 54.2±23.1%, respectively. The total recovery of method B was the highest, and that of method C was higher than that of method A which was the lowest (P<0.05). The recoveries (mean±SD) of various types of diatoms were 2.95 ±4.11%,96.26±120.50% and 66.74±99.17% in the three groups, respectively. The diatom species recovery of method B was the highest, and that of method C was higher than that of method A which was the lowest (P<0.05).Case test:The relative rate of diatom species was 39.8±34.6% in the lung of drowning group with method A, and the number of diatoms were 4936.3±9863/10g. The species and the number of diatoms were lower than those in method B and those in method C in the liver and renal of the drowning groups. The relative rate of diatom species was 69.6±38.5% in the lung of drowning group with method B, and the number of diatom were 52743.5±82018.8/10g. The species and the number of diatoms were the higest in the liver and renal of the drowning groups with method B.The relative rate of diatom species was 39.8±34.6% in the lung of drowning group with method A, and the number of diatoms were 4936.3±9863/10g. The species and the number of diatoms were higher than those in method A in the liver and renal of the drowning groups. The positive rate of method A was between diatom Method B test positive rate between methods C diatom test positive rate between. The sampling time of method A was the longest, and the sampling time of method B was longer than that of method C, whose time was the shortest (P<0.05). The diatoms in the liver and kidney were difficult to observe under the methods A and the diatoms numbers were detected few. The diatoms in the lung, liver and kidney were easy observed under the method B. The diatoms in the lung and water sample were difficult to observe under the method C because there were many silicon particles hinder the diatom identification.Conclusion:This study created a new method of diatom test which based on membrane enrichment and optical microscopy. Diatom in the digestion solution were enriched in the PES membrane, and employed on the eugenol and acetic acid mixed reagent, so that the membrane could be observed directly under an optical microscope. Samples in three groups corpses (drowned corpses, bodies thrown into water after death and the bodies dead in the land) were imposed on microwave digestion-membrane enrichment-light microscopy, and also compared the results of those of imposed on other two common diatom test. This method characters high sensitivity, ability to avoid contamination, and simple procedure, efficiency, less cost, easy to learn, etc. It was suitable for forensic science laboratory. |
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