Detection of diverse aquatic microbes in blood and organs of drowning victims: First metagenomic approach using high-throughput 454-pyrosequencing

Kakizaki, Eiji; Ogura, Yoshitoshi; Kozawa, Shuji; Nishida, Sho; Uchiyama, Taketo; Hayashi, Tetsuya; Yukawa, Nobuhiro

doi:10.1016/j.forsciint.2012.02.010

Cited by 54 publications

(25 citation statements)

References 30 publications

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“…Microorganisms can be detected by incubation in selective media or by various PCR techniques. In addition to DNA fragments specific for some aquatic bacteria, the 16S ribosomal RNA gene (16 rDNA) has been repeatedly used as a target for bacterial and planktonic elements (He et al 2008;Tie et al 2010;Huys et al 2012;Kakizaki et al 2012). …”

Section: Microbiological Testsmentioning

confidence: 99%

Injuries due to Asphyxiation and Drowning

Keil¹,

Lunetta²,

Vann³

et al. 2014

Handbook of Forensic Medicine

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Section: Microbiological Testsmentioning

confidence: 99%

Injuries due to Asphyxiation and Drowning

Keil¹,

Lunetta²,

Vann³

et al. 2014

Handbook of Forensic Medicine

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“…Prior approaches to bacterial identification by DNA sequencing have included fluorescent Sanger sequencing of hundreds of small variable regions of 16S ribosomal amplimer clones (19,20), length heterogeneity (LH) PCR for 16S ribosomal variable regions (21), restriction fragment polymorphisms in 16S (19,21), arrays with bacterium-specific probes, such as PhyloChip (22,23), and early applications of NGS (24,25). These methods have been applied to various clinical situations, such as the microbiome of suboptimally controlled asthma (22), cystic fibrosis (19,21), chronic obstructive pulmonary disease (COPD) (24,26,27), bronchiectasis (28), pleurisy (20), forensic assessment of drowning (25), and intubated patients (23). These studies have revealed unexpectedly large numbers of bacterial genera in patients with clinical illnesses and in healthy individuals (29,30).…”

mentioning

confidence: 99%

Single-Molecule Long-Read 16S Sequencing To Characterize the Lung Microbiome from Mechanically Ventilated Patients with Suspected Pneumonia

et al. 2014

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iIn critically ill patients, the development of pneumonia results in significant morbidity and mortality and additional health care costs. The accurate and rapid identification of the microbial pathogens in patients with pulmonary infections might lead to targeted antimicrobial therapy with potentially fewer adverse effects and lower costs. Major advances in next-generation sequencing (NGS) allow culture-independent identification of pathogens. The present study used NGS of essentially full-length PCRamplified 16S ribosomal DNA from the bronchial aspirates of intubated patients with suspected pneumonia. The results from 61 patients demonstrated that sufficient DNA was obtained from 72% of samples, 44% of which (27 samples) yielded PCR amplimers suitable for NGS. Out of the 27 sequenced samples, only 20 had bacterial culture growth, while the microbiological and NGS identification of bacteria coincided in 17 (85%) of these samples. Despite the lack of bacterial growth in 7 samples that yielded amplimers and were sequenced, the NGS identified a number of bacterial species in these samples. Overall, a significant diversity of bacterial species was identified from the same genus as the predominant cultured pathogens. The numbers of NGSidentifiable bacterial genera were consistently higher than identified by standard microbiological methods. As technical advances reduce the processing and sequencing times, NGS-based methods will ultimately be able to provide clinicians with rapid, precise, culture-independent identification of bacterial, fungal, and viral pathogens and their antimicrobial sensitivity profiles.

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“…Evidence suggests that other species of bacteria present in water are translocated into the organs during drowning [10]. In cases where bodies are found in water where the species targeted by this PCR are absent, other bacterial species could be used to provide evidence for the cause of death.…”

Section: Limitations Of Studymentioning

confidence: 99%

“…In cases where bodies are found in water where the species targeted by this PCR are absent, other bacterial species could be used to provide evidence for the cause of death. A more broad-based approach, such as microbiome profiling using next-generation sequencing [10], although likely to have its own limitations, has the potential to overcome this disadvantage of targeted PCR.…”

Section: Limitations Of Studymentioning

confidence: 99%

“…Although their work appeared promising, doubts still existed in relation to the use of faecal bacteria for the diagnosis of drowning [9]. Following on from this early work, a series of papers has been presented within the peer reviewed literature from Japan concerning the investigation of bacterioplankton, first in blood and latterly in tissue samples for the diagnosis of drowning [10][11][12][13][14][15][16][17]. This work has shown that using molecular approaches, most recently real-time PCR assays with TaqMan probes for bacterioplankton, it is possible to undertake rapid, less laborious, high throughput tests to differentiate freshwater from marine bacterioplankton and in doing so provide a molecular diagnostic test to assist in the diagnosis of drowning [18].…”

Section: Introductionmentioning

confidence: 99%

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Detection of bacterioplankton using PCR probes as a diagnostic indicator for drowning; the Leicester experience

Rutty¹,

Bradley²,

Biggs³

et al. 2015

Legal Medicine

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Bodies found immersed in water can pose difficulties to the investigating authorities. Pathologists may be assisted with the diagnosis by the use of tests such as the analysis for diatoms or the levels of strontium in the blood, although there is a recognised level of uncertainty associated with these tests. Recent work from Japan has shown that using molecular approaches, most recently real-time polymerase chain reaction (PCR) assays with TaqMan probes for bacterioplankton, it is possible to undertake rapid, less laborious, high throughput tests to differentiate freshwater from marine bacterioplankton and in doing so provide a molecular diagnostic test to assist in the diagnosis of drowning. We report the experiences of a United Kingdom forensic pathology unit in the use of this PCR based system for the diagnosis of drowning. We applied this technique to 20 adult and child cadavers from 4 bath, 12 freshwater, 2 brackish and 2 salt water scenes both from within the United Kingdom and abroad. Drowning was concluded to be the cause of death in 16 of these 20 cases and the PCR method supported this conclusion in 12 of these 16 cases. The PCR did not provide evidence of drowning in the four cases where death was from other causes. We illustrate that this PCR method provides a rapid diagnostic supportive test for the diagnosis of drowning that can be applied to United Kingdom autopsy practice.

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Detection of diverse aquatic microbes in blood and organs of drowning victims: First metagenomic approach using high-throughput 454-pyrosequencing

Cited by 54 publications

References 30 publications

Injuries due to Asphyxiation and Drowning

Injuries due to Asphyxiation and Drowning

Single-Molecule Long-Read 16S Sequencing To Characterize the Lung Microbiome from Mechanically Ventilated Patients with Suspected Pneumonia

Detection of bacterioplankton using PCR probes as a diagnostic indicator for drowning; the Leicester experience

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