2007 Fiscal Year Final Research Report Summary
Molecular analysis of anteliotic-tolerance and stress response in adherent bacteria
| Project/Area Number |
17591914
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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 |
Morphological basic dentistry
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| Research Institution | The University of Tokushima |
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Principal Investigator |
ONO Tsuneko The University of Tokushima, School of Medicine, Professor (40035514)
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| Co-Investigator(Kenkyū-buntansha) |
MIYAKE Yoichiro The University of Tokushima, Institute of Health Biosciences, Graduate School, Professor (80136093)
ABIKO Yoshimitsu Nihon University, School of Dentistry at Matsudo, Professor (70050086)
HIGUCHI Tomihiko The University of Tokushima, Institute of Health Biosciences, Graduate School, professor (50035557)
KUWAHARA Tomomi The University of Tokushima, Institute of Health Biosciences, Graduate School, Associate Professor (60263810)
KAYAMA Shizuo The University of Tokushima, Institute of Health Biosciences, Graduate School, Assistant Professor (50432761)
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| Project Period (FY) |
2005 – 2007
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| Keywords | antibiotic tolerance / stress response / rpoS / adherent bacteria / ppGppc / rpoN / pvdS / vqsR |
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| Research Abstract |
In this project, we have studied about the molecular mechanism of antibiotic tolerance and stress response in adherent cells The following results werte obtained in this study.
1. To assess the contribution of ppGpp in antibiotic tolerance to quinolone in Pseudomonas aeruginosa, knockout mutants of relA, spoT and dksA, were constructed and investigated for their antibiotic susceptibility to quinolones. The survival of the dksA and spoT mutants in the presence of ofloxacin and ciprofloxacin were shown to be approximately 20-180 and 10-40 times respectively, higher than the same for the wild type strain.
The data suggest that elevated basal levels of ppGpp may be responsible for rendering these mutants tolerant to quinolones and expand the importance of ppGpp as an antimicrobial target in P. aeruginosa
2. An rpoN mutant in Pseudomonas aeruginosa has been constructed and investigated its importance as a target for antibiotics. The stationary phase cells of the rpoN mutant displayed an approximately 15 times higher survival rate to quinolones and carbapenems than the wild-type cells. Real-time PCR revealed that in contrast to the wild-type strain, the stationary phase cells of the rpoN mutant grown without antibiotics had approximately 4-140 and 2-14-fold higher transcripts of pvdS and vqsR genes, respectively, than the wild type strain. In the presence of antibiotics, pvdS and vqsR transcripts were 400 and 5-fold, respectively, elevated in comparison to the wild-type. Flow cytometry assays using GFP reporter demonstrated increased expression of the vqsR gene in the rpoN mutant. A pvdS mutant of P. aeruginosa, deficient in pyoverdine production, was shown to be susceptible to biapenem.
These findings suggest that rpoN is involved in the tolerance to antimicrobial agents in P. aeruginosa and its tolerant effect is partly dependent on increased pyoverdine production and vqsR gene expression.
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