| Project/Area Number |
25420562
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Research Field |
Civil and environmental engineering
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| Research Institution | Hachinohe National College of Technology |
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Principal Investigator |
Yaguchi Junichi 八戸工業高等専門学校, その他部局等, 教授 (80342450)
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| Co-Investigator(Kenkyū-buntansha) |
Yamamoto Ayumu 八戸工業高等専門学校, 産業システム工学科マテリアル・バイオコース, 准教授 (60523800)
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| Co-Investigator(Renkei-kenkyūsha) |
Kaneko Nakaichiro 八戸工業高等専門学校, 産業システム工学科環境都市・建築デザインコース, 助手 (70099761)
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| Project Period (FY) |
2013-04-01 – 2016-03-31
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| Project Status |
Completed (Fiscal Year 2015)
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| Budget Amount *help |
¥4,940,000 (Direct Cost: ¥3,800,000、Indirect Cost: ¥1,140,000)
Fiscal Year 2015: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2014: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2013: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
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| Keywords | 大腸菌 / DNA / LED / PMA試薬 / 青色LED / LED光源 / VBNC状態 |
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| Outline of Final Research Achievements |
Using membrane filtration, we first developed and optimized a protocol for sample treatment with propidium monoazide (PMA) in combination with real-time PCR(PMA-qPCR) to distinguish viable and dead Escherichia coli cells. Water sample treatment with 50 μM PMA for 5 min followed by 5min of halogen light exposure resulted in an effective exclusion of DNA from heat-treated E.coli cells. When applied to water environmental samples, the viable E.coli counts based on the PMA-qPCR method were almost intermediate between the total and culturable counts in water environments. A number of E.coli remained in the VBNC state and the proportion of VBNC cells to total ones ranged from 2.3 to 87.9%.
In addition, light emitting diode (LED) was used to avoid excessive heating of samples as an alternative to halogen light. 15 and 40 min of LED light exposure time were sufficient to exclude DNA from heat-treated E. coli cells in suspension and on membrane filter, respectively.
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