1988 Fiscal Year Final Research Report Summary
抗アラリア剤ピリメサミンの標的酵素遺伝子のクローニングとその薬剤耐性機構の解明
| Project/Area Number |
62570175
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
寄生虫学(含医用動物学）
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| Research Institution | Okayama University Medical School |
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Principal Investigator |
ISHII Akira Dept. Parasitol., Prof., 医学部, 教授 (40012752)
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| Co-Investigator(Kenkyū-buntansha) |
WATAYA Yusuke Fuculty of Pharmaceutical Science. Ass.Prof., 薬学部, 助教授 (90127598)
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| Project Period (FY) |
1987 – 1988
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| Keywords | Malaria / Pyrimethamine / Drug resistance / ELISA / 倍養 |
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| Research Abstract |
Malaria parasite, (Plasmodium falciparum) used were FCR-3(Gumma Univ.), FUP(Osaka Univ.) and FCO-1(Okayama Univ.,isolated in North Sumatra) strains. These were adapted to CO2 incubator culture. We tried to make FCR-3 strain resistant to pyrimethamine, methotrexate and trimethoprim by adding increasing doses step by step. They became to show a certain degree of resistance to drugs, however there was a lop of difficulties in attaining high degree of resistance.
We made and assembled an equipment to run pulse field electrophoresis (OFAGE). We run chromosomal large sized DNA of malaria parasites. We could not detect any difference among the strains especially that of gene amplification.
We measured drug susceptibility of the malaria strains. Results were obtained by classical culture method, however it took more than 3 days for us to obtain drug resistance information and needed time consuming microscopic examination. We tried to use anti-BrdU antibody in ELISA system to measure incorporation of bromodeoxyuridine into the parasites. The system was established as a novel short time assay for drug susceptibility testing. The method was proved useful in the field in North Sumatra.
Resistance to pyrimethamine was reported to be caused by a point mutation in dehydrofolate reductase gene. We tried to prepare the probe to detect not only malaria parasite but also mutation of the gene. The fundamental experimentaions are under way.
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