Studies on the method of conserving fungal strains of medical importance
| Project/Area Number |
60480168
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
細菌学
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| Research Institution | Teikyo University |
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Principal Investigator |
YAMAGUCHI Hideyo Research Institute for Medical Mycology, Teikyo University, 医学部, 教授 (40009890)
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| Co-Investigator(Kenkyū-buntansha) |
MIYAJI Makoto Research Institute of Chemobiodynamics, Chiba University, 生物活性研究所, 教授 (40009494)
UCHIDA Katsuhisa Research Institute for Medical Mycology, Teikyo University, 医真菌研究センター, 講師 (00009995)
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| Project Period (FY) |
1985 – 1986
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| Project Status |
Completed (Fiscal Year 1986)
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| Budget Amount *help |
¥6,200,000 (Direct Cost: ¥6,200,000)
Fiscal Year 1986: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1985: ¥5,100,000 (Direct Cost: ¥5,100,000)
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| Keywords | pathogenic fungi / conservation of living cultures / programmed-freezing method / programmed freezer / 凍結乾燥法 |
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| Research Abstract |
For the purpose of establishing theprogrammed freezing and freeze-drying as the useful method for maintenance and preservation of pathogenic fungi for a long period, suitable conditions for processing fungal cultures in these two methods were investigated by using 39 strains of 22 fungal species as the testing organism. The appropriateness of the conditions chosen for preserving living fungal cultures was caaessed on the basis of comparison of viable counts, morphological, physiological, serological and/or genetic characteristics, and pathogenicity or virulence to mice of the cultures that was carried out before and after freezing or freeze-drying processes.
A large number or experiments have been done in this laboratory with the programmed freezing method and appreciable success was achieved when the following procedures were employed. In our procedure, fresh cultures of fungi (especially yeasts) are grown on Sabouraud glucose agar in test tubes. A heavy suspension of vegetative cells
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or spores (approx. <10^8> organisms/ml) is prepared in sterile 10%(w/v) dimethyl sulfoxide(DMSO), and approximately 1 ml of the suspension is added to each of several sterile,capped plastic vials. The filled vials are directly placed into the freezing chamber of a programmed freezer. The initial cooling is carried out slowly at a rate of 1 to 2゜C/min from room temperature to -45゜C; subsequent cooling at a more rapid rate of 5 to 6゜C/ min to -85゜C. Then the vials are immediately transferred to storage in liquid nitrogen(LN). All strains of pathogenic fungi are stored in the vapor phase of an LN refrigerator and handled with care. For recovery of the fungal cultures frozen in LN, the frozen vials are thawed rapidly in 35゜C water bath until the last trace of ice is dissipated. Under these experimental conditions, 31 to 100% of the viable units are usually recovered. There are variance in the recovery rate of fungal cultures among different strain within the same species. Any significant changes in the morphological, physiological and/or serological properties, as well as pathogenicity (or virulence) to mice, of fungal cultures were not produced by the processes of freezing or subsequent thawing, as compared with untreated comparable cultures. It was also demonstrated that the recovery rate of cultures preserved for 6 to 12 months according to the above-mentioned programmed freezing procedures was significatly greater than that of corresponding cultures which had been subjected to slow freezing by transfer into a deep freezer. These results suggest the crucial importance of temperature-controlled freezing and, therefore, usefulness of the programmed freezer for successful conservation of pathogenic fungi. Less
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Report
(1 results)
Research Products
(6 results)
