2002 Fiscal Year Final Research Report Summary
A novel PCR method for identifying plankton in cases of death by drowning
| Project/Area Number |
13670430
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Legal medicine
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| Research Institution | FUKUSHIMA MEDICAL UNIVERSITY |
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Principal Investigator |
HIRAIWA Kouichi Fukushima Med. Univ., Schl. Of Med., Professor, 医学部, 教授 (60124616)
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| Co-Investigator(Kenkyū-buntansha) |
KURISAKI Emiko Fukushima Med. Univ., Schl. Of Med., Assistant Professor, 医学部, 講師 (30106356)
GUNJI Hirobumi Fukushima Med. Univ., Schl. Of Med., Assistant Professor, 医学部, 講師 (20234643)
ABE Sumiko Fukushima Med. Univ., Schl. Of Med., Research Assistant, 医学部, 助手 (50136975)
MIZUSAWA Ikumumi Fukushima Med. Univ., Schl. Of Med., Research Assistant, 医学部, 助手 (40192356)
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| Project Period (FY) |
2001 – 2002
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| Keywords | Plankton / PCR / Chlorophyll-related sequences / drowning |
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| Research Abstract |
We developed a new PCR method for identifying plankton in cases of death by drowning. We designed 4 primer pairs for chlorophyll-related genes of Euglena gracilis (EG) and Skeletonema costatum (SK), which are commonly distributed in water. The primers were selected from sequences coding chloroplast/chlorophyll apoprotein of EG (EG1 and EG2) and fucoxanthin-chlorophyll a/c harvesting protein of SK (SK1 and SK2). With EG1 or EG2, up to 2 fg of EG-DNA was identified, and 0.2 pg of SK-DNA was detectable with SK1 or SK2. No PCR products were amplified from green vegetables (komatsuna, spinach, parsley) or human DNA with the 4 primer pairs. Regardless of origin, seawater or fresh water, most diatoms were detectable with primer pairs of EG1 and EG2. With SK1, only Centrales diatoms were identified, and 5 diatom strains originating from seawater were detectable with SK2. EG1 and EG2 gave rise to PCR products from most water samples. By using Percoll, plankton was easily isolated from human tissue and blood samples with which various water samples obtained from the sea, a river and a dam had been mixed. Plankton genes were detected in lung, liver and kidney homogenates mixed with 1 to 3 ml of water samples. By using Percoll, plankton was easily isolated from human tissue or blood samples, and good results of PCR analysis were obtained in cases of death by drowning. PCR products were amplified from lung, liver and kidney samples obtained from drowning victims that showed diatom-negative results using the acid digestion method. In order to evaluate the usefulness of the plankton method for diagnosis of drowning, we examined plankton genes in a non-drowned rabbit submerged after death. Plankton genes were detected in lung samples but not in heart blood samples. Because of postmortem plankton invasion into lungs, lungs are not suitable for plankton identification.
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