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The Journal of the Japanese Society for Clinical Microbiology |
Biblioraphy Information
| ArticleTitle |
Construction of Inspection System for M. pneumoniae Rapid Detection Method in Outpatients, Using Real-Time PCR Method |
| Language |
J |
| AuthorList |
Akihiro Nakamura, Noriyuki Abe, Hisashi Kono, Yoshiko Fujimoto, Saori Fukuda, Noriko Hatanaka, Syuji Matsuo |
| Affiliation |
Department of Clinical Bacteriology, Clinical Pathology, Tenri Hospital |
| Publication |
J.J.C.M.: 19 (3), 148-156, 2009 |
| Received |
December 26, 2008 |
| Accepted |
May 11, 2009 |
| Abstract |
Atypical pneumonia pathogens hold about 10 to 30% among community-acquired pneumonia pathogens in Japan. The quick and accurate detection of atypical pneumonia pathogens, such as M. pneumoniae, is difficult by the routine outpatient-based laboratory inspections. We established the M. pneumoniae rapid detection method in outpatients, using real-time PCR method. For the purpose of establishing the inspection methods, we focused on three points as follows. First of all we used SYBR Green I as a fluorescence detection format and 2.0 Light Cycler system, which enabled us to make up a 45-minutes rapid detection method from specimen presentation to report of results. The primer on M. pneumoniae 16S rRNA gene was used for this PCR method, and sensitivity for M. pneumoniae in the minimum was showed to be 3.09 fg/μl, which is equivalent to about 102-103 copies in number. It seems that we could be able to sufficiently cope with an acute phase of M. pneumoniae pneumonia, although in which period numbers of bacteria could be abundant. Next, we tried to omit the DNA extraction used in common genetic diagnostic examinations. We compared crossing point values between two situations, one is performing DNA extraction and the other is not, in 15 cases diagnosed as M. pneumoniae pneumonia. As a result, the PCR sensitivity did not show any estrangement in both methods. Finally we examined about PCR inhibitors. Antibacterial agents did not influenced the PCR process, but blood did lower the sensitivity of the PCR. We believe that introduction of this genetic methodology into routine bacteriological examinations, such as Gram staining, bacteriological culture tests and serological examination, can contribute to quicker and accurate bacteriological diagnosis. |
| Keywords |
real-time PCR, Mycoplasma pneumoniae, 16S rRNA gene |
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