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首页> 外文期刊>Forensic science international >A new molecular approach to help conclude drowning as a cause of death: Simultaneous detection of eight bacterioplankton species using real-time PCR assays with TaqMan probes
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A new molecular approach to help conclude drowning as a cause of death: Simultaneous detection of eight bacterioplankton species using real-time PCR assays with TaqMan probes

机译：一种新的分子方法可以帮助将溺水归结为死亡原因：使用TaqMan探针的实时PCR检测法同时检测八种浮游细菌

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We developed a novel tool for concluding drowning as a cause of death. We designed nine primer pairs to detect representative freshwater or marine bacterioplankton (aquatic bacteria) and then used real-time PCR with TaqMan probes to rapidly and specifically detect them. We previously cultured the genus Aeromonas, which is a representative freshwater bacterial species, in blood samples from 94% of victims who drowned in freshwater and the genera Vibrio and/or Photobacterium that are representative marine bacteria in 88% of victims who drowned in seawater. Based on these results, we simultaneously detected eight species of bacterioplankton (Aeromonas hydrophila, A. salmonicida; Vibrio fischeri, V. harveyi, V. parahaemolyticus; Photobacterium damselae, P. leiognathi, P. phosphoreum) using three sets of triplex real-time PCR assays and TaqMan probes labelled with fluorophores (FAM, NED, Cy5). We assayed 266 specimens (109 blood, 157 tissues) from 43 victims, including 32 who had drowned in rivers, ditches, wells, sea or around estuaries. All lung samples of these 32 victims were TaqMan PCR-positive including the lung periphery into which water does not readily enter postmortem. On the other hand, findings in blood and/or closed organs (kidney or liver) were PCR-positive in 84% of the drowned victims (except for those who drowned in baths) although the conventional test detected diatoms in closed organs in only 44% of the victims. Thus, the results of the PCR assay reinforced those of diatom tests when only a few diatoms were detectable in organs due to the low density of diatoms in the water where they were found. Multiplex TaqMan PCR assays for bacterioplankton were rapid, less laborious and high-throughput as well as sensitive and specific. Therefore, these assays would be useful for routine forensic screening tests to estimate the amount and type of aspirated water.
机译：我们开发了一种新颖的工具来将溺水归为死亡原因。我们设计了九对引物来检测代表性的淡水或海洋浮游细菌（水生细菌），然后使用TaqMan探针进行实时PCR快速，特异地检测它们。我们以前在94％的淡水溺水受害者和88％的海水溺水受害者中代表海洋细菌的弧菌和/或光细菌属的血液样本中培养了代表典型淡水细菌物种的气单胞菌属。根据这些结果，我们使用三套实时三套实时检测方法同时检测了八种浮游细菌（嗜水气单胞菌，鲑鱼气单胞菌;费氏弧菌，哈氏弧菌，副溶血性弧菌;水母光子菌，白僵菌，磷杆菌）。 PCR检测和TaqMan探针标记有荧光团（FAM，NED，Cy5）。我们分析了43位受害者的266个标本（109个血液，157个组织），其中包括32位淹没在河流，沟渠，水井，海洋或河口附近的人。这32位受害者的所有肺部样本均为TaqMan PCR阳性，包括肺部周围的水分不容易进入死后。另一方面，在84％的溺水受害者中，血液和/或封闭器官（肾脏或肝脏）中的PCR阳性（除了那些在浴池中溺水的患者），尽管常规检测仅在44个器官中检测到了硅藻在封闭器官中受害者百分比。因此，当在器官中只能检测到少量硅藻时，由于在水中发现的硅藻密度低，因此PCR分析的结果增强了硅藻测试的结果。 TaqMan PCR检测浮游生物的快速，省力，高通量以及灵敏性和特异性。因此，这些测定对于常规的法医筛查测试以估计所吸入的水的量和类型将是有用的。

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