1998 Fiscal Year Final Research Report Summary
Detection of gene mutations in kanamycin-resistant Mycobacterium tuberculosis and development of the method for rapid diagnosis
| Project/Area Number |
08670470
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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 |
Public health/Health science
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| Research Institution | Osaka Prefectural Institute of Public Health |
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Principal Investigator |
KATSUKAWA Chihiro Osaka Prefectural Institute of Public Health, Department of Public Health, Senior Research Scientist, 公衆衛生部, 主任研究員 (20183725)
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| Co-Investigator(Kenkyū-buntansha) |
TAMARU Aki Osaka Prefectural Institute of Public Health, Department of Public Health, Resea, 公衆衛生部, 研究員 (70270767)
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| Project Period (FY) |
1996 – 1998
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| Keywords | Mycobacterium tuberculosis / kanamycin / kanamycin-resistant strains / rrs gene / PCR-RFLP analysis |
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| Research Abstract |
In Mycobacterium smegmatis and a limited number of Mycobacterium tuberculosis strains, the involvement of alterations of the 16S rRNA gene (rrs) in resistance to kanamycin has been shown. To investigate the extent to which mutations in a specific region of the n-s gene and the kanamycin-resistant phenotype in clinically isolated M.tuberculosis swains were correlated, 43 kanamycin-resistant swains (MICs, >=200 mu g/ml), 71 kanamycin-susceptible strains, and 4 type strains were examined. The 300-bp DNA fragments carrying the rrs gene and the intervening sequence between the rrs gene and 23S rRNA (rrl) gene fragments were amplified by PCR and were subjected to PCR-based direct sequencing. By comparing the nucleotide sequences, substitutions were found in 29 of 43 (67.4%) kanamycin-resistant clinical isolates at position 1400, 1401 and 1483 but in none of the 71 sensitive isolates or the 4 type strains. The most frequent substitution, from A to G, occurred at position 1400. A substitution from C to T at position 1401 was found once. Two clinical isolates carried the double mutation from C to A at position 1401 and from G to T at position 1483. In addition, we found that these mutations can be distinguished from wild-type strains by digestion with the restriction endonucleases Tai I and Tsp 451. Furthermore, we found that the genotypes of kanamycin-resistant swains can be discriminated from each other by digestion with a restriction endonuclease, Bst UI or Dde I.
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