| Project/Area Number |
07041162
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| Research Category |
Grant-in-Aid for international Scientific Research
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| Allocation Type | Single-year Grants |
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| Section | Field Research |
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
AMAKO Kazunobu Kyushu University, Professor, 医学部, 教授 (20078752)
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| Co-Investigator(Kenkyū-buntansha) |
ISLAM M.sirajul ICDDR,B.Associate Scientist, B. Environmental Microbiology, Associate
MORIYA Tetsuhiro Kyushu University, Research Associate, 医学部, 助手 (10140790)
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| Project Period (FY) |
1995
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| Project Status |
Completed (Fiscal Year 1995)
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| Budget Amount *help |
¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1995: ¥1,000,000 (Direct Cost: ¥1,000,000)
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| Keywords | Vibrio cholerae / Nonculturable / Reactivation / Heat-shock / Ammonium ion |
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| Research Abstract |
We have established a viable but nonculturable state of Vibrio cholerae strain TSI-4 by incubating in M9 salt or phosphate-buffered saline (PBS) at 15ﾟC.After 25-35 days the bacteria lost the colony forming ability on nutrient agar plate and the shape of the bacteria transferred to coccoid form. The ATP concentration in the cells was, however, kept at a constant level throughout experimental period at 15ﾟC.The recovery of viable cells from this stage was achieved when the culture was heated at 45ﾟC for one min. The nonculturable cells established in PBS,however, was not resuscitated by heat shock treatment. Since the difference of M9 salt and PBS was the presence of ammonium salt in M9 salt but not in PBS,we carried out the experiment to recover the bacteria from PBS supplemented with NH_4Cl and we had the colonies on agar plates after heat shock from this culture. The recovered cells were confirmed to be identical to the bacteria that originally used in this experiment. This reactivation method was applied to V.cholerae in environmental water samples conllected in Bangladesh. Though the water samples did not contain any cultivable V.cholerae and any detectable PCR product of ctx gene, from some samples that was incubated at 45ﾟC in the presence of ammonium salt we could detect ctx gene after amplification by PCR.We also had colonies of V.cholerae in such heated samples and identified them as O1 Inaba serogroup of V.cholerae.
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